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Su W, Yang P, Xu F, Zhang T, Wang L, Li H, Cui L, Yang Z, He H, Han S, He L, Liu J, Kong Y, Long J. Twin Strep-Tag Modified CPT1A Mitochondrial Membrane Chromatography in Screening Lipid Metabolism Regulators. Anal Chem 2024; 96:10851-10859. [PMID: 38912707 DOI: 10.1021/acs.analchem.4c02402] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 06/25/2024]
Abstract
Mitochondrial Membrane Chromatography (MMC) is a bioaffinity chromatography technique developed to study the interaction between target proteins embedded in the mitochondrial membrane and their ligand compounds. However, the MMC stationary phases (MMSP) prepared by chemical immobilization are prone to nonspecific binding in candidate agent screening inevitably. To address these challenges, Twin Strep-Tag/Strep Tactin was employed to establish a specific affinity system in the present study. We prepared a carnitine palmitoyltransferase 1A (CPT1A) MMSP by specifically linking Strep-tactin-modified silica gel with the Twin Strep-Tag on the CPT1A-oriented mitochondrial membrane. This Twin Strep-Tag/Strep Tactin modified CPT1A/MMC method exhibited remarkably better retention behavior, longer stationary phase lifespan, and higher screening specificity compared with previous MMC systems with glutaraldehyde immobilization. We adopted the CPT1A-specific MMC system in screening CPT1A ligands from traditional Chinese medicines, and successfully identified novel candidate ligands: ononin, isoliquiritigenin, and aloe-emodin, from Glycyrrhiza uralensis Fisch and Senna tora (L.) Roxb extracts. Biological assessments illustrated that the compounds screened promote CPT1A enzyme activity without affecting CPT1A protein expression, as well as effectively reduce the lipid droplets and triglyceride levels in the high fat induction HepG2 cells. The results suggest that we have developed an MMC system, which is promising for studying the bioaffinity of mitochondrial membrane proteins to candidate compounds. This system provides a platform for a key step in mitochondrial medicine discovery, especially for bioactive molecule screening from complex herbal extracts.
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Affiliation(s)
- Wu Su
- Center for Mitochondrial Biology and Medicine, The Key Laboratory of Biomedical Information Engineering of Ministry of Education, School of Life Science and Technology, Xi'an Jiaotong University, Xi'an 710049, China
| | - Peng Yang
- Center for Mitochondrial Biology and Medicine, The Key Laboratory of Biomedical Information Engineering of Ministry of Education, School of Life Science and Technology, Xi'an Jiaotong University, Xi'an 710049, China
| | - Fanding Xu
- Center for Mitochondrial Biology and Medicine, The Key Laboratory of Biomedical Information Engineering of Ministry of Education, School of Life Science and Technology, Xi'an Jiaotong University, Xi'an 710049, China
| | - Tingrong Zhang
- Center for Mitochondrial Biology and Medicine, The Key Laboratory of Biomedical Information Engineering of Ministry of Education, School of Life Science and Technology, Xi'an Jiaotong University, Xi'an 710049, China
| | - Lizhuo Wang
- Center for Mitochondrial Biology and Medicine, The Key Laboratory of Biomedical Information Engineering of Ministry of Education, School of Life Science and Technology, Xi'an Jiaotong University, Xi'an 710049, China
| | - Hua Li
- Center for Mitochondrial Biology and Medicine, The Key Laboratory of Biomedical Information Engineering of Ministry of Education, School of Life Science and Technology, Xi'an Jiaotong University, Xi'an 710049, China
| | - Li Cui
- Center for Mitochondrial Biology and Medicine, The Key Laboratory of Biomedical Information Engineering of Ministry of Education, School of Life Science and Technology, Xi'an Jiaotong University, Xi'an 710049, China
| | - Zhiwei Yang
- School of Physics, Xi'an Jiaotong University, Xi'an 710116, China
| | - Huaizhen He
- School of Pharmacy, Xi'an Jiaotong University, Xi'an 710116, China
| | - Shengli Han
- School of Pharmacy, Xi'an Jiaotong University, Xi'an 710116, China
| | - Langchong He
- School of Pharmacy, Xi'an Jiaotong University, Xi'an 710116, China
| | - Jiankang Liu
- Center for Mitochondrial Biology and Medicine, The Key Laboratory of Biomedical Information Engineering of Ministry of Education, School of Life Science and Technology, Xi'an Jiaotong University, Xi'an 710049, China
| | - Yu Kong
- Center for Mitochondrial Biology and Medicine, The Key Laboratory of Biomedical Information Engineering of Ministry of Education, School of Life Science and Technology, Xi'an Jiaotong University, Xi'an 710049, China
| | - Jiangang Long
- Center for Mitochondrial Biology and Medicine, The Key Laboratory of Biomedical Information Engineering of Ministry of Education, School of Life Science and Technology, Xi'an Jiaotong University, Xi'an 710049, China
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Su W, Xu F, Zhong J, Hu R, Wang L, Li H, Yang Z, Ge S, He H, Han S, Xie X, Guo H, He L, Liu J, Yi T, Kong Y, Long J. Screening of CPT1A-Targeting Lipid Metabolism Modulators Using Mitochondrial Membrane Chromatography. ACS APPLIED MATERIALS & INTERFACES 2024; 16:13234-13246. [PMID: 38411590 DOI: 10.1021/acsami.3c18102] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 02/28/2024]
Abstract
Carnitine palmitoyltransferase 1A (CPT1A), which resides on the mitochondrial outer membrane, serves as the rate-limiting enzyme of fatty acid β-oxidation. Identifying the compounds targeting CPT1A warrants a promising candidate for modulating lipid metabolism. In this study, we developed a CPT1A-overexpressed mitochondrial membrane chromatography (MMC) to screen the compounds with affinity for CPT1A. Cells overexpressing CPT1A were cultured, and subsequently, their mitochondrial membrane was isolated and immobilized on amino-silica gel cross-linked by glutaraldehyde. After packing the mitochondrial membrane column, retention components of MMC were performed with LC/MS, whose analytic peaks provided structural information on compounds that might interact with mitochondrial membrane proteins. With the newly developed MMC-LC/MS approach, several Chinese traditional medicine extracts, such as Scutellariae Radix and Polygoni Cuspidati Rhizoma et Radix (PCRR), were analyzed. Five noteworthy compounds, baicalin, baicalein, wogonoside, wogonin, and resveratrol, were identified as enhancers of CPT1A enzyme activity, with resveratrol being a new agonist for CPT1A. The study suggests that MMC serves as a reliable screening system for efficiently identifying modulators targeting CPT1A from complex extracts.
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Affiliation(s)
- Wu Su
- Center for Mitochondrial Biology and Medicine, the Key Laboratory of Biomedical Information Engineering of Ministry of Education, School of Life Science and Technology, Xi'an Jiaotong University, Xi'an 710049, China
| | - Fanding Xu
- Center for Mitochondrial Biology and Medicine, the Key Laboratory of Biomedical Information Engineering of Ministry of Education, School of Life Science and Technology, Xi'an Jiaotong University, Xi'an 710049, China
| | - Jinjin Zhong
- Center for Mitochondrial Biology and Medicine, the Key Laboratory of Biomedical Information Engineering of Ministry of Education, School of Life Science and Technology, Xi'an Jiaotong University, Xi'an 710049, China
| | - Ranrui Hu
- Center for Mitochondrial Biology and Medicine, the Key Laboratory of Biomedical Information Engineering of Ministry of Education, School of Life Science and Technology, Xi'an Jiaotong University, Xi'an 710049, China
| | - Lizhuo Wang
- Center for Mitochondrial Biology and Medicine, the Key Laboratory of Biomedical Information Engineering of Ministry of Education, School of Life Science and Technology, Xi'an Jiaotong University, Xi'an 710049, China
| | - Hua Li
- Center for Mitochondrial Biology and Medicine, the Key Laboratory of Biomedical Information Engineering of Ministry of Education, School of Life Science and Technology, Xi'an Jiaotong University, Xi'an 710049, China
| | - Zhiwei Yang
- School of Physics, Xi'an Jiaotong University, Xi'an 710116, China
| | - Shuai Ge
- School of Pharmacy, Xi'an Jiaotong University, Xi'an 710116, China
| | - Huaizhen He
- School of Pharmacy, Xi'an Jiaotong University, Xi'an 710116, China
| | - Shengli Han
- School of Pharmacy, Xi'an Jiaotong University, Xi'an 710116, China
| | - Xiuying Xie
- Department of Gynecology and Obstetrics, Second Affiliated Hospital of Medical College, Xi'an Jiaotong University, Xi'an 710005, China
| | - Hui Guo
- Department of Endocrinology, First Affiliated Hospital of Medical College, Xi'an Jiaotong University, Xi'an 710000, China
| | - Langchong He
- School of Pharmacy, Xi'an Jiaotong University, Xi'an 710116, China
| | - Jiankang Liu
- School of Health and Life Sciences, University of Health and Rehabilitation Sciences, Qingdao, Shandong 266113, China
- Department of Dermatology of the First Affiliated Hospital, Xi'an Jiaotong University, Xi'an 710049, China
| | - Tao Yi
- Faculty of Health Sciences and Sports, Macao Polytechnic University, Macau 999078, China
| | - Yu Kong
- Center for Mitochondrial Biology and Medicine, the Key Laboratory of Biomedical Information Engineering of Ministry of Education, School of Life Science and Technology, Xi'an Jiaotong University, Xi'an 710049, China
| | - Jiangang Long
- Center for Mitochondrial Biology and Medicine, the Key Laboratory of Biomedical Information Engineering of Ministry of Education, School of Life Science and Technology, Xi'an Jiaotong University, Xi'an 710049, China
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Yun H, Liu Z, Hou W, Liu Q, Nong Y, Li S, Liu C. Rapid screening and isolation of 5-lipoxygenase inhibitors in Inonotus obliquus and mechanism of action in the treatment of asthma. J Sep Sci 2024; 47:e2300647. [PMID: 38466162 DOI: 10.1002/jssc.202300647] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/05/2023] [Revised: 01/02/2024] [Accepted: 01/31/2024] [Indexed: 03/12/2024]
Abstract
Accurate screening and targeted preparative isolation of active substances in natural medicines have long been two technical challenges in natural medicine research. This study outlines a new approach to improve the efficiency of natural product preparation, focusing on rapidly and accurately screening potential active ingredients in Inonotus obliquus as well as efficiently preparing 5-lipoxidase (5-LOX) inhibitors, to provide new ideas for the treatment of asthma with Inonotus obliquus. First, we used ultrafiltration (UF) mass spectrometry to screen for three potential inhibitors of 5-LOX in Inonotus obliquus. Subsequently, the inhibitory effect of the active ingredients screened in the UF assay on 5-LOX was verified using the molecular docking technique, and the potential role of the active compounds in Inonotus obliquus for the treatment of asthma was analyzed by network pharmacology. Finally, based on the above activity screening guidelines, we used semi-preparative liquid chromatography and consecutive high-speed countercurrent chromatography to isolate three high-purity 5-LOX inhibitors such as betulin, lanosterol, and quercetin. Obviously, through the above approach, we have seamlessly combined rapid discovery, screening, and centralized preparation of the active ingredient with molecular-level interactions between the active ingredient and the protease.
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Affiliation(s)
- Haocheng Yun
- Central Laboratory, Changchun Normal University, Changchun, Jilin, P. R. China
| | - Zhen Liu
- Central Laboratory, Changchun Normal University, Changchun, Jilin, P. R. China
| | - Wanchao Hou
- Central Laboratory, Changchun Normal University, Changchun, Jilin, P. R. China
| | - Qiang Liu
- Central Laboratory, Changchun Normal University, Changchun, Jilin, P. R. China
| | - Yuyu Nong
- Central Laboratory, Changchun Normal University, Changchun, Jilin, P. R. China
| | - Sainan Li
- Central Laboratory, Changchun Normal University, Changchun, Jilin, P. R. China
| | - Chunming Liu
- Central Laboratory, Changchun Normal University, Changchun, Jilin, P. R. China
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Li Z, Li B, Liu M, Chen Z, Li P, Du R, Su M, Anirudhan V, Achi JG, Tian J, Rong L, Cui Q. Development of a virus-based affinity ultrafiltration method for screening virus-surface-protein-targeted compounds from complex matrixes: Herbal medicines as a case study. J Med Virol 2024; 96:e29517. [PMID: 38476091 DOI: 10.1002/jmv.29517] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/18/2023] [Revised: 02/13/2024] [Accepted: 02/27/2024] [Indexed: 03/14/2024]
Abstract
Herbal medicines (HMs) are one of the main sources for the development of lead antiviral compounds. However, due to the complex composition of HMs, the screening of active compounds within these is inefficient and requires a significant time investment. We report a novel and efficient virus-based screening method for antiviral active compounds in HMs. This method involves the centrifugal ultrafiltration of viruses, known as the virus-based affinity ultrafiltration method (VAUM). This method is suitable to identify virus specific active compounds from complex matrices such as HMs. The effectiveness of the VAUM was evaluated using influenza A virus (IAV) H1N1. Using this method, four compounds that bind to the surface protein of H1N1 were identified from dried fruits of Terminalia chebula (TC). Through competitive inhibition assays, the influenza surface protein, neuraminidase (NA), was identified as the target protein of these four TC-derived compounds. Three compounds were identified by high performance liquid chromatography (HPLC) and liquid chromatography/mass spectrometry (LC/MS), and their anti-H1N1 activities were verified by examining the cytopathic effect (CPE) and by performing a virus yield reduction assay. Further mechanistic studies demonstrated that these three compounds directly bind to NA and inhibit its activity. In summary, we describe here a VAUM that we designed, one that can be used to accurately screen antiviral active compounds in HMs and also help improve the efficiency of screening antiviral drugs found in natural products.
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Affiliation(s)
- Zhongyuan Li
- College of Pharmacy, Shandong University of Traditional Chinese Medicine, Jinan, China
| | - Baohong Li
- Innovative Institute of Chinse Medicine and Pharmacy, Shandong University of Traditional Chinese Medicine, Jinan, China
| | - Miaomiao Liu
- College of Pharmacy, Shandong University of Traditional Chinese Medicine, Jinan, China
| | - Zinuo Chen
- Innovative Institute of Chinse Medicine and Pharmacy, Shandong University of Traditional Chinese Medicine, Jinan, China
| | - Ping Li
- Key Laboratory of Chemical Biology (Ministry of Education), Department of Medicinal Chemistry, School of Pharmaceutical Sciences, Shandong University, Jinan, China
| | - Ruikun Du
- Innovative Institute of Chinse Medicine and Pharmacy, Shandong University of Traditional Chinese Medicine, Jinan, China
- Qingdao Academy of Chinese Medicinal Sciences, Shandong University of Traditional Chinese Medicine, Qingdao, China
| | - Ming Su
- Shandong Academy of Chinese Medicine, Jinan, China
| | - Varada Anirudhan
- Department of Microbiology and Immunology, College of Medicine, University of Illinois Chicago, Chicago, Illinois, USA
| | - Jazmin G Achi
- Department of Microbiology and Immunology, College of Medicine, University of Illinois Chicago, Chicago, Illinois, USA
| | - Jingzhen Tian
- College of Pharmacy, Shandong University of Traditional Chinese Medicine, Jinan, China
- Qingdao Academy of Chinese Medicinal Sciences, Shandong University of Traditional Chinese Medicine, Qingdao, China
| | - Lijun Rong
- Department of Microbiology and Immunology, College of Medicine, University of Illinois Chicago, Chicago, Illinois, USA
| | - Qinghua Cui
- Innovative Institute of Chinse Medicine and Pharmacy, Shandong University of Traditional Chinese Medicine, Jinan, China
- Qingdao Academy of Chinese Medicinal Sciences, Shandong University of Traditional Chinese Medicine, Qingdao, China
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Guo Y, Gong P, Qian Y, Liu H, Yu B, Qi J. Rapid screening and identification of superoxide dismutase activators from traditional Chinese medicines based on affinity ultrafiltration mass chromatography combined with molecular docking. J Chromatogr A 2023; 1710:464408. [PMID: 37804579 DOI: 10.1016/j.chroma.2023.464408] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/21/2023] [Revised: 09/20/2023] [Accepted: 09/20/2023] [Indexed: 10/09/2023]
Abstract
In the present study, a comprehensive strategy integrating affinity ultrafiltration high-performance liquid chromatography quadrupole-time-of-flight mass spectrometry (UF-HPLC-Q-TOF-MS), in silico molecular docking and bioassays was established to rapidly screen natural SOD activators from traditional Chinese medicines. As illustrative case studies, Schisandra chinensis, Fructus cnidii and Radix ophiopogonis were chosen to develop and verify the strategy. The HPLC-Q-TOF-MS was used to identify the compounds in comparison with reference standards and literature data. A total of eight compounds, including four biphenyl-cyclooctene ligands from Schisandra chinensis and four coumarins from Fructus cnidii, were found to potentially increase SOD activities. No ligands were found in the extract of Radix ophiopogonis. Then, in silico molecular docking was performed to investigate the binding site and binding affinity of the candidates on SOD. Compared to the nonspecific ligands screened from the extract, the specific ligands presented stronger binding affinities. In addition, the activity and kinetic parameters of the SOD-ligand were investigated through an improved pyrogallol autoxidation method. Gomisin J and xanthotoxin showed a stronger ability to increase SOD activities. The present study indicated that combining UF-HPLC-Q-TOF-MS and in silico molecular docking offers a powerful and meaningful tool to rapidly screen SOD activators from traditional Chinese medicines.
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Affiliation(s)
- Yujie Guo
- Jiangsu Key Laboratory of TCM Evaluation and Translational Research, School of Traditional Chinese Pharmacy, China Pharmaceutical University, Nanjing 211198, PR China; School of Pharmacy, Hubei University of Chinese Medicine, Wuhan 430065, PR China
| | - Puyang Gong
- Jiangsu Key Laboratory of TCM Evaluation and Translational Research, School of Traditional Chinese Pharmacy, China Pharmaceutical University, Nanjing 211198, PR China
| | - Yin Qian
- Jiangsu Key Laboratory of TCM Evaluation and Translational Research, School of Traditional Chinese Pharmacy, China Pharmaceutical University, Nanjing 211198, PR China
| | - Haichun Liu
- Jiangsu Key Laboratory of TCM Evaluation and Translational Research, School of Traditional Chinese Pharmacy, China Pharmaceutical University, Nanjing 211198, PR China
| | - Boyang Yu
- Jiangsu Key Laboratory of TCM Evaluation and Translational Research, School of Traditional Chinese Pharmacy, China Pharmaceutical University, Nanjing 211198, PR China.
| | - Jin Qi
- Jiangsu Key Laboratory of TCM Evaluation and Translational Research, School of Traditional Chinese Pharmacy, China Pharmaceutical University, Nanjing 211198, PR China.
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Wang T, Li YQ, Yu LP, Zi L, Yang YQ, Zhang M, Hao JJ, Gu W, Zhang F, Yu J, Yang XX. Compatibility of Polygonati Rhizoma and Angelicae Sinensis Radix enhance the alleviation of metabolic dysfunction-associated fatty liver disease by promoting fatty acid β-oxidation. Biomed Pharmacother 2023; 162:114584. [PMID: 36989710 DOI: 10.1016/j.biopha.2023.114584] [Citation(s) in RCA: 2] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/26/2023] [Revised: 03/08/2023] [Accepted: 03/21/2023] [Indexed: 03/30/2023] Open
Abstract
Jiuzhuan Huangjing Pills (JHP) composed of Polygonati Rhizoma (PR) and Angelicae Sinensis Radix (ASR) remedied mitochondria to cure metabolic dysfunction-associated fatty liver disease (MAFLD). However, a comparison of the anti-MAFLD ability between JHP prescription and PR and ASR single-medicines in MAFLD has not been performed, and the action mechanisms and substances remain unknown. Our results show that JHP, PR and ASR decreased serum and liver lipid levels. The effects of JHP were stronger than those of PR and ASR. JHP, PR and ASR afforded protection to mitochondrial ultrastructure, and regulated oxidative stress and energy metabolism in mitochondria. JHP also regulated the expression of β-oxidation genes, which were not regulated by PR and ASR. JHP-, PR- and ASR-derived components in mitochondrial extracts regulated oxidative stress, energy metabolism, and β-oxidation gene expression and alleviated cellular steatosis. Four, six and eleven compounds were identified in mitochondrial extracts from PR-, ASR- and JHP-treated rats, respectively. The data suggest that JHP, PR and ASR alleviated MAFLD by remedying mitochondria, while the ability of JHP was stronger than that of PR and ASR, which was involved with the β-oxidation promotion. The compounds identified may be the main ingredients in the three extracts active in MAFLD improvement.
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Han L, Wang H, Cao J, Li Y, Jin X, He C, Wang M. Inhibition mechanism of α-glucosidase inhibitors screened from Tartary buckwheat and synergistic effect with acarbose. Food Chem 2023; 420:136102. [PMID: 37060666 DOI: 10.1016/j.foodchem.2023.136102] [Citation(s) in RCA: 21] [Impact Index Per Article: 10.5] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/02/2022] [Revised: 03/01/2023] [Accepted: 03/31/2023] [Indexed: 04/17/2023]
Abstract
Tartary buckwheat has been shown to provide a good antihyperglycemic effect. However, it is unclear which active compounds play a key role in attenuating postprandial hyperglycemia. Presently, acetone extract from the hull of Tartary buckwheat had the best effect for α-glucosidase inhibition (IC50 = 0.02 mg/mL). Twelve potential α-glucosidase inhibitors from Tartary buckwheat were screened and identified by the combination of ultrafiltration and high-performance liquid chromatography coupled with mass spectrometry. Myricetin and quercetin exhibited the highest anti-α-glucosidase activity with IC50 values of 0.02 and 0.06 mg/mL, respectively. These inhibitors manifested different types of inhibition manners against α-glucosidase via direct interaction with the amino acid residues. The results of structure-activity relationships indicated that an increase in the number of -OH on the B-ring greatly strengthened α-glucosidase inhibitory activity, but glucoside and rutinoside replacement on the C-ring obviously weakened this influence. Furthermore, a synergistic effect was observed between inhibitors with different inhibition manners.
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Affiliation(s)
- Lin Han
- College of Food Science and Engineering, Northwest A&F University, Yangling 712100, PR China
| | - Huiqing Wang
- College of Food Science and Engineering, Northwest A&F University, Yangling 712100, PR China
| | - Junwei Cao
- College of Food Science and Engineering, Northwest A&F University, Yangling 712100, PR China
| | - Yunlong Li
- Institute of Functional Food of Shanxi, Shanxi Agricultural University, Taiyuan 030006, PR China
| | - Xiying Jin
- College of Food Science and Engineering, Northwest A&F University, Yangling 712100, PR China
| | - Caian He
- College of Food Science and Engineering, Northwest A&F University, Yangling 712100, PR China.
| | - Min Wang
- College of Food Science and Engineering, Northwest A&F University, Yangling 712100, PR China.
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Yu LP, Li YJ, Wang T, Tao YX, Zhang M, Gu W, Yu J, Yang XX. In vivo recognition of bioactive substances of Polygonum multiflorum for protecting mitochondria against metabolic dysfunction-associated fatty liver disease. World J Gastroenterol 2023; 29:171-189. [PMID: 36683716 PMCID: PMC9850952 DOI: 10.3748/wjg.v29.i1.171] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 09/07/2022] [Revised: 11/01/2022] [Accepted: 12/05/2022] [Indexed: 01/04/2023] Open
Abstract
BACKGROUND Metabolic dysfunction-associated fatty liver disease (MAFLD) is a severe threat to human health. Polygonum multiflorum (PM) has been proven to remedy mitochondria and relieve MAFLD, but the main pharmacodynamic ingredients for mitigating MAFLD remain unclear. AIM To research the active ingredients of PM adjusting mitochondria to relieve high-fat diet (HFD)-induced MAFLD in rats. METHODS Fat emulsion-induced L02 adipocyte model and HFD-induced MAFLD rat model were used to investigate the anti-MAFLD ability of PM and explore their action mechanisms. The adipocyte model was also applied to evaluate the activities of PM-derived constituents in liver mitochondria from HFD-fed rats (mitochondrial pharmacology). PM-derived constituents in liver mitochondria were confirmed by ultra-high-performance liquid chromatography/mass spectrometry (mitochondrial pharmacochemistry). The abilities of PM-derived monomer and monomer groups were evaluated by the adipocyte model and MAFLD mouse model, respectively. RESULTS PM repaired mitochondrial ultrastructure and prevented oxidative stress and energy production disorder of liver mitochondria to mitigate fat emulsion-induced cellular steatosis and HFD-induced MAFLD. PM-derived constituents that entered the liver mitochondria inhibited oxidative stress damage and improved energy production against cellular steatosis. Eight chemicals were found in the liver mitochondria of PM-administrated rats. The anti-steatosis ability of one monomer and the anti-MAFLD activity of the monomer group were validated. CONCLUSION PM restored mitochondrial structure and function and alleviated MAFLD, which may be associated with the remedy of oxidative stress and energy production. The identified eight chemicals may be the main bioactive ingredients in PM that adjusted mitochondria to prevent MAFLD. Thus, PM provides a new approach to prevent MAFLD-related mitochondrial dysfunction. Mitochondrial pharmacology and pharmacochemistry further showed efficient strategies for determining the bioactive ingredients of Chinese medicines that adjust mitochondria to prevent diseases.
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Affiliation(s)
- Li-Ping Yu
- College of Pharmaceutical Science, Yunnan University of Chinese Medicine, Kunming 650500, Yunnan Province, China
- College of Pharmaceutical Science, Yunnan Key Laboratory of Southern Medicine Utilization, Kunming 650500, Yunnan Province, China
| | - Yan-Juan Li
- College of Pharmaceutical Science, Yunnan University of Chinese Medicine, Kunming 650500, Yunnan Province, China
- College of Pharmaceutical Science, Yunnan Key Laboratory of Southern Medicine Utilization, Kunming 650500, Yunnan Province, China
| | - Tao Wang
- College of Pharmaceutical Science, Yunnan University of Chinese Medicine, Kunming 650500, Yunnan Province, China
- College of Pharmaceutical Science, Yunnan Key Laboratory of Southern Medicine Utilization, Kunming 650500, Yunnan Province, China
| | - Yu-Xuan Tao
- College of Pharmaceutical Science, Yunnan University of Chinese Medicine, Kunming 650500, Yunnan Province, China
- College of Pharmaceutical Science, Yunnan Key Laboratory of Southern Medicine Utilization, Kunming 650500, Yunnan Province, China
| | - Mei Zhang
- College of Pharmaceutical Science, Yunnan University of Chinese Medicine, Kunming 650500, Yunnan Province, China
- College of Pharmaceutical Science, Yunnan Key Laboratory of Southern Medicine Utilization, Kunming 650500, Yunnan Province, China
| | - Wen Gu
- College of Pharmaceutical Science, Yunnan University of Chinese Medicine, Kunming 650500, Yunnan Province, China
- College of Pharmaceutical Science, Yunnan Key Laboratory of Southern Medicine Utilization, Kunming 650500, Yunnan Province, China
| | - Jie Yu
- College of Pharmaceutical Science, Yunnan University of Chinese Medicine, Kunming 650500, Yunnan Province, China
- College of Pharmaceutical Science, Yunnan Key Laboratory of Southern Medicine Utilization, Kunming 650500, Yunnan Province, China
| | - Xing-Xin Yang
- College of Pharmaceutical Science, Yunnan University of Chinese Medicine, Kunming 650500, Yunnan Province, China
- College of Pharmaceutical Science, Yunnan Key Laboratory of Southern Medicine Utilization, Kunming 650500, Yunnan Province, China
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In vivo identification of the pharmacodynamic ingredients of Polygonum cuspidatum for remedying the mitochondria to alleviate metabolic dysfunction–associated fatty liver disease. Biomed Pharmacother 2022; 156:113849. [DOI: 10.1016/j.biopha.2022.113849] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/07/2022] [Revised: 09/08/2022] [Accepted: 10/06/2022] [Indexed: 11/21/2022] Open
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10
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Tang N, Hong F, Hao W, Yu TT, Wang GG, Li W. Riboflavin ameliorates mitochondrial dysfunction via the AMPK/PGC1α/HO‑1 signaling pathway and attenuates carbon tetrachloride‑induced liver fibrosis in rats. Exp Ther Med 2022; 24:608. [PMID: 36160891 PMCID: PMC9468838 DOI: 10.3892/etm.2022.11545] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/08/2022] [Accepted: 07/06/2022] [Indexed: 12/01/2022] Open
Abstract
Hepatic fibrosis is a global health problem, with increasing evidence demonstrating that oxidative stress serves a pivotal role in fibrogenesis. Riboflavin is a vital nutrient in the human and animal diet, which enhances the activity of antioxidant enzymes and ameliorates oxidative stress. The present study evaluated the effect of riboflavin on liver fibrosis and the mechanisms underlying this process. Rats were subcutaneously injected with carbon tetrachloride (CCl4) dissolved in sterile olive oil twice per week to induce hepatic fibrosis. The effect of riboflavin on CCl4-induced liver fibrosis was then assessed. Blood samples and liver tissues were collected and analyzed. The liver tissue morphological changes, immunohistochemical analysis, levels of malondialdehyde (MDA) and superoxide dismutase (SOD) in the mitochondria, and the protein expression levels of α-smooth muscle actin (α-SMA), transforming growth factor-β1 (TGF-β1), extracellular signal-regulated kinase (ERK), p38, c-Jun N-terminal kinase (JNK), AMP-activated protein kinase (AMPK), peroxisome proliferator-activated receptor γ coactivator-1α (PGC-1α) and heme oxygenase 1 (HO-1) in the liver were also analyzed. The results demonstrated that riboflavin treatment significantly decreased the levels of alanine transaminase and aspartate transaminase in the serum, increased SOD activity and modulated the MDA level in the mitochondria. Furthermore, riboflavin significantly inhibited the CCl4-induced, upregulated protein expression levels of phosphorylated (p)-ERK, p-p38, p-JNK, TGF-β1 and α-SMA. Moreover, riboflavin significantly increased the expression of p-AMPK, PGC-1α and HO-1 in the liver tissue. These results suggested that riboflavin delays CCl4-induced hepatic fibrosis by enhancing the mitochondrial function via the AMPK/PGC-1α/HO-1 and mitogen-activated protein kinase signaling pathways.
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Affiliation(s)
- Ning Tang
- Emergency Intensive Care Unit, The First Affiliated Hospital of Wannan Medical College, Wuhu, Anhui 241002, P.R. China
| | - Feng Hong
- Department of Physiology, Wannan Medical College, Wuhu, Anhui 241002, P.R. China
| | - Wei Hao
- Department of Experimental Center for Function Subjects, Wannan Medical College, Wuhu, Anhui 241002, P.R. China
| | - Ting-Ting Yu
- Department of Experimental Center for Function Subjects, Wannan Medical College, Wuhu, Anhui 241002, P.R. China
| | - Guo-Guang Wang
- Department of Pathophysiology, Wannan Medical College, Wuhu, Anhui 241002, P.R. China
| | - Wei Li
- Department of Pathophysiology, Wannan Medical College, Wuhu, Anhui 241002, P.R. China
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11
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Zhang F, Li L, Zhao JH, Ge XT, Gao H, Jia TZ. The Effects of Salt-Water Processing of Phellodendri Chinensis Cortex on the Enhancement of Kidney Absorption of the Main Alkaloids. Nat Prod Commun 2022. [DOI: 10.1177/1934578x221076218] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/15/2022] Open
Abstract
Phellodendri chinensis cortex (PC) is a commonly used traditional Chinese medicine for clearing heat. It is often applied in the clinical practice by using the salt-water processing method. According to the theory of Chinese meteria medica processing, after being processed with salt-water, traditional Chinese medicine can increase absorption of the kidney and enhance the effects of kidney disease treatments, which can be abbreviated as “entering into kidney by processing with salt-water”. PC is a typical traditional Chinese medicine using the salt-water processing method. The resulting salt-water processed PC（SPC）can specifically enter the kidney meridian, as compared to raw PC (RPC), which enhances the kidney yin nourishing and purges away kidney fire. However, the effect of “entering into kidney by processing with salt-water” of PC has not been proven and its mechanism is unknown. Therefore, it is of great significance to compare the renal absorption effect of PC before and after salt-water processing and elucidate its mechanism. In this article, using the PC alkaloid content as an indicator, a human proximal tubular epithelial cell (HK-2 cell) experiment and the binding rate of alkaloids to lysozyme-kidney target enzyme was was conducted to simulate kidney absorption in vitro. These results were combined with an investigation of the alkaloid content in rat kidneys after a gavage of PC solution, comprehensively evaluating the difference in kidney absorption after salt-water processing. Compared with RPC, the efflux of SPC alkaloids in the HK-2 cell permeability experiment was significantly reduced, and affinity of the lysozyme-kidney target enzyme was higher. In addition, those who were given SPC had a higher alkaloid concentration in the kidneys than the RPC group. Finally, we verified enhanced kidney absorption effect of PC by salt-water processing, and to a certain extent revealed the mechanism of enhanced kidney absorption of SPC.
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Affiliation(s)
- Fan Zhang
- School of Pharmacy, Liaoning University of Traditional Chinese Medicine, Dalian, China
| | - Li Li
- School of Pharmacy, Liaoning University of Traditional Chinese Medicine, Dalian, China
| | - Jia-hui Zhao
- School of Pharmacy, Liaoning University of Traditional Chinese Medicine, Dalian, China
| | - Xiu-tong Ge
- School of Pharmacy, Liaoning University of Traditional Chinese Medicine, Dalian, China
| | - Hui Gao
- School of Pharmacy, Liaoning University of Traditional Chinese Medicine, Dalian, China
| | - Tian-zhu Jia
- School of Pharmacy, Liaoning University of Traditional Chinese Medicine, Dalian, China
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12
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A Novel Method for Identifying Parkin Binding Agents in Complex Preparations of Herbal Medicines. OXIDATIVE MEDICINE AND CELLULAR LONGEVITY 2022; 2022:3260243. [PMID: 35087614 PMCID: PMC8789414 DOI: 10.1155/2022/3260243] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 08/23/2021] [Revised: 11/11/2021] [Accepted: 12/04/2021] [Indexed: 11/24/2022]
Abstract
Parkin is a crucial E3 ubiquitin ligase for initiating mitophagy through the PINK1/Parkin pathway. Regulating the expression and activity of parkin can remedy mitophagy and human disease. We developed an efficient method to isolate natural parkin ligands from herbal medicines by combining centrifugal ultrafiltration and liquid chromatography/mass spectrometry. The heterologous expression technology identified functionally active and pure parkin proteins. After evaluating the reliability of the method using DL-selenomethionine and DL-dithiothreitol as positive controls, this method was successfully applied to capture parkin ligands from Polygoni Cuspidati Rhizoma et Radix and Sophorae Flavescentis Radix. LC/MS identified seven novel parkin-targeting compounds, namely, 7,4′-dihydroxy-5-methoxy-8-(γ, γ-dimethylallyl)-flavanone, kushenol I, kurarinone, sophoraflavanone G, torachrysone-8-O-glucoside, apigenin, and emodin, supported by the molecular docking analysis. Five of the seven novel compounds (kushenol I, kurarinone, sophoraflavanone G, apigenin, and emodin) can activate parkin in in vitro autoubiquitination assays. Meanwhile, kushenol I and kurarinone had antisteatosis activity in fat emulsion-damaged human hepatocytes. These results confirmed the effectiveness of the method for identifying parkin ligands from complex preparations, useful to advance drug discovery from medicinal herbs.
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13
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Muchiri RN, van Breemen RB. Drug discovery from natural products using affinity selection-mass spectrometry. DRUG DISCOVERY TODAY. TECHNOLOGIES 2021; 40:59-63. [PMID: 34916024 DOI: 10.1016/j.ddtec.2021.10.005] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Subscribe] [Scholar Register] [Received: 09/24/2020] [Revised: 10/08/2021] [Accepted: 10/11/2021] [Indexed: 12/22/2022]
Abstract
As a starting point for drug discovery, affinity selection-mass spectrometry (AS-MS) is ideal for the discovery of lead compounds from chemically diverse sources such as botanical, fungal and microbial extracts. Based on binding interactions between macromolecular receptors and ligands of low molecular mass, AS-MS enables the rapid isolation of pharmacologically active small molecules from complex mixtures for mass spectrometric characterization and identification. Unlike conventional high-throughput screening, AS-MS requires no radiolabels, no UV or fluorescent chromophores, and is compatible with all classes of receptors, enzymes, incubation buffers, cofactors, and ligands. The most successful types of AS-MS include pulsed ultrafiltration (PUF) AS-MS, size exclusion chromatography (SEC) AS-MS, and magnetic microbead affinity selection screening (MagMASS), which differ in their approaches for separating the ligand-receptor complexes from the non-binding compounds in mixtures. After affinity isolation, the ligand(s) from the mixture are characterized using high resolution UHPLC-MS and tandem mass spectrometry. Based on these elemental composition and structural data, the identities of the lead compounds are determined by searching on-line databases for known natural products and by comparison with standards. The structures of novel natural products are determined using a combination of spectroscopic techniques including two-dimensional NMR and MS.
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Affiliation(s)
- Ruth N Muchiri
- Linus Pauling Institute, Oregon State University, Corvallis, OR 97331, United States; College of Pharmacy, Oregon State University, Corvallis, OR 97331, United States
| | - Richard B van Breemen
- Linus Pauling Institute, Oregon State University, Corvallis, OR 97331, United States; College of Pharmacy, Oregon State University, Corvallis, OR 97331, United States
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14
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Identification and characterization of the human urinary metabolites of trimetazidine using liquid chromatography high resolution mass spectrometry, an anti-doping perspective. Microchem J 2021. [DOI: 10.1016/j.microc.2021.106872] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/22/2022]
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15
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Mu JK, Zi L, Li YQ, Yu LP, Cui ZG, Shi TT, Zhang F, Gu W, Hao JJ, Yu J, Yang XX. Jiuzhuan Huangjing Pills relieve mitochondrial dysfunction and attenuate high-fat diet-induced metabolic dysfunction-associated fatty liver disease. Biomed Pharmacother 2021; 142:112092. [PMID: 34449316 DOI: 10.1016/j.biopha.2021.112092] [Citation(s) in RCA: 7] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/22/2021] [Revised: 08/14/2021] [Accepted: 08/19/2021] [Indexed: 01/03/2023] Open
Abstract
BACKGROUND Metabolic dysfunction-associated fatty liver disease (MAFLD) is a common global chronic liver disease. Jiuzhuan Huangjing Pills (JHP) have been used for the treatment of human disease for over a thousand years, but their efficacy and underlying mechanism(s) of action against MAFLD are unknown. We investigated the alleviating effects of JHP on high-fat diet (HFD)-induced MAFLD. METHODS In vitro and in vivo methods were used to evaluate the effects of JHP on MAFLD. L02 adipocyte models were induced by fat emulsion and adipocytes were treated with JHP for 24 h. MAFLD rat models were induced by HFD-feeding and were intragastrically administered JHP for 12 weeks. Changes in fat accumulation, L02 cell damage, body weight, food intake, histological parameters, organ indexes, biochemical parameters, and mitochondrial indicators including ultrastructure, oxidative stress, energy metabolism, and fatty acid metabolism were investigated. RESULTS JHP attenuated the increase in levels of total cholesterol, triglyceride, low density lipoprotein cholesterol, alanine transaminase, and aspartate transaminase levels, and significantly increased high density lipoprotein cholesterol. JHP up-regulated levels of glutathione (GSH) and superoxide dismutase (SOD), and down-regulated malondialdehyde (MDA). JHP afforded protection to the mitochondrial ultrastructure, and inhibited the HFD-induced increase in MDA and the reduction of SOD, GSH, ATP synthase, and complex I and II, in liver mitochondria. JHP regulated the expression of β-oxidation genes, including acyl-CoA dehydrogenase, cyl-CoA dehydrogenase long chain, carnitine palmitoyltransferase 1A, carnitine palmitoyltransferase 1B, peroxisomal proliferator-activated receptor-gamma coactivator-1α and peroxide proliferator activated receptor α. CONCLUSION JHP alleviates HFD-induced MAFLD through the protection of mitochondrial function.
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Affiliation(s)
- Jian-Kang Mu
- College of Pharmaceutical Science, Yunnan University of Chinese Medicine, 1076 Yuhua Road, Kunming 650500, China; Yunnan Key Laboratory of Southern Medicine Utilization, 1076 Yuhua Road, Kunming 650500, China
| | - Lei Zi
- College of Pharmaceutical Science, Yunnan University of Chinese Medicine, 1076 Yuhua Road, Kunming 650500, China; Yunnan Key Laboratory of Southern Medicine Utilization, 1076 Yuhua Road, Kunming 650500, China
| | - Yan-Qin Li
- College of Pharmaceutical Science, Yunnan University of Chinese Medicine, 1076 Yuhua Road, Kunming 650500, China; Yunnan Key Laboratory of Southern Medicine Utilization, 1076 Yuhua Road, Kunming 650500, China
| | - Li-Ping Yu
- College of Pharmaceutical Science, Yunnan University of Chinese Medicine, 1076 Yuhua Road, Kunming 650500, China; Yunnan Key Laboratory of Southern Medicine Utilization, 1076 Yuhua Road, Kunming 650500, China
| | - Zheng-Guo Cui
- Department of Environmental Health, University of Fukui School of Medical Science, 23-3 Matsuoka Shimoaizuki, Eiheiji 910-1193, Japan
| | - Ting-Ting Shi
- Department of Pharmaceutical Preparation, The Xixi Hospital of Hangzhou Affiliated to Zhejiang University of Traditional Chinese Medicine, Hangzhou 310023, China
| | - Fan Zhang
- College of Pharmaceutical Science, Yunnan University of Chinese Medicine, 1076 Yuhua Road, Kunming 650500, China; Yunnan Key Laboratory of Southern Medicine Utilization, 1076 Yuhua Road, Kunming 650500, China
| | - Wen Gu
- College of Pharmaceutical Science, Yunnan University of Chinese Medicine, 1076 Yuhua Road, Kunming 650500, China; Yunnan Key Laboratory of Southern Medicine Utilization, 1076 Yuhua Road, Kunming 650500, China
| | - Jun-Jie Hao
- College of Pharmaceutical Science, Yunnan University of Chinese Medicine, 1076 Yuhua Road, Kunming 650500, China; Yunnan Key Laboratory of Southern Medicine Utilization, 1076 Yuhua Road, Kunming 650500, China
| | - Jie Yu
- College of Pharmaceutical Science, Yunnan University of Chinese Medicine, 1076 Yuhua Road, Kunming 650500, China; Yunnan Key Laboratory of Southern Medicine Utilization, 1076 Yuhua Road, Kunming 650500, China.
| | - Xing-Xin Yang
- College of Pharmaceutical Science, Yunnan University of Chinese Medicine, 1076 Yuhua Road, Kunming 650500, China; Yunnan Key Laboratory of Southern Medicine Utilization, 1076 Yuhua Road, Kunming 650500, China.
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16
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Yang J, Wang W, Qiang C, Niu Y, Li Z, Zhao M, Jiang Y, Cao J, Niu X, Liu X, Zhao J. Effects of a fully enclosed hollow-fiber centrifugal ultrafiltration technique for laboratory biosafety improvement. Biotechniques 2021; 71:465-472. [PMID: 34350779 DOI: 10.2144/btn-2021-0029] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/23/2022] Open
Abstract
Laboratory biosafety has become a core focus in biological analysis, owing to the frequent occurrence of laboratory-acquired infections caused by the leakage of pathogenic microorganisms. For this purpose, the authors developed a safe pretreatment device combining a sealing technique with a direct injection technique. In this study, several bacteria and viruses were used to validate the filtration effect of the invention. Data show that the new device can completely filter bacteria and that the filtration rates for hepatitis B virus and hepatitis C virus reached 94% and 96%, respectively. The results show that the new preparation device can effectively block these pathogens and can improve biological safety and provide powerful protection for technicians.
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Affiliation(s)
- Jing Yang
- Hebei Provincial Center for Clinical Laboratories, Second Hospital of Hebei Medical University, 215 West Heping Road, Shijiazhuang, Hebei Province, 050051, China
| | - Weigang Wang
- Hebei Provincial Center for Clinical Laboratories, Second Hospital of Hebei Medical University, 215 West Heping Road, Shijiazhuang, Hebei Province, 050051, China
| | - Cuixin Qiang
- Hebei Provincial Center for Clinical Laboratories, Second Hospital of Hebei Medical University, 215 West Heping Road, Shijiazhuang, Hebei Province, 050051, China
| | - Yanan Niu
- Hebei Provincial Center for Clinical Laboratories, Second Hospital of Hebei Medical University, 215 West Heping Road, Shijiazhuang, Hebei Province, 050051, China
| | - Zhirong Li
- Hebei Provincial Center for Clinical Laboratories, Second Hospital of Hebei Medical University, 215 West Heping Road, Shijiazhuang, Hebei Province, 050051, China
| | - Mengqiang Zhao
- Department of Pharmaceutical Analysis, School of Pharmacy, Hebei Medical University, 361 Zhongshan East Road, Shijiazhuang, Hebei Province, 050000, China
| | - Ye Jiang
- Department of Pharmaceutical Analysis, School of Pharmacy, Hebei Medical University, 361 Zhongshan East Road, Shijiazhuang, Hebei Province, 050000, China
| | - Jing Cao
- Hebei Provincial Center for Clinical Laboratories, Second Hospital of Hebei Medical University, 215 West Heping Road, Shijiazhuang, Hebei Province, 050051, China
| | - Xiaoran Niu
- Hebei Provincial Center for Clinical Laboratories, Second Hospital of Hebei Medical University, 215 West Heping Road, Shijiazhuang, Hebei Province, 050051, China
| | - Xiaoxuan Liu
- Hebei Provincial Center for Clinical Laboratories, Second Hospital of Hebei Medical University, 215 West Heping Road, Shijiazhuang, Hebei Province, 050051, China
| | - Jianhong Zhao
- Hebei Provincial Center for Clinical Laboratories, Second Hospital of Hebei Medical University, 215 West Heping Road, Shijiazhuang, Hebei Province, 050051, China
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17
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da Cunha Menezes Souza L, Chen M, Ikeno Y, Salvadori DMF, Bai Y. The implications of mitochondria in doxorubicin treatment of cancer in the context of traditional and modern medicine. TRADITIONAL MEDICINE AND MODERN MEDICINE 2021. [DOI: 10.1142/s2575900020300076] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/18/2022] Open
Abstract
Doxorubicin (DOX) is an antibiotic anthracycline extensively used in the treatment of different malignancies, such as breast cancer, lymphomas and leukemias. The cardiotoxicity induced by DOX is one of the most important pathophysiological events that limit its clinical application. Accumulating evidence highlights mitochondria as a central role in this process. Modulation of mitochondrial functions as therapeutic strategy for DOX-induced cardiotoxicity has thus attracted much attention. In particular, emerging studies investigated the potential of natural mitochondria-targeting compounds from Traditional Chinese Medicine (TCM) as adjunct or alternative treatment for DOX-induced toxicity. This review summarizes studies about the mechanisms of DOX-induced cardiotoxicity, evidencing the importance of mitochondria and presenting TCM treatment alternatives for DOX-induced cardiomyopathy.
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Affiliation(s)
| | - Meng Chen
- School of Traditional Chinese Medicine, Beijing University of Traditional Chinese Medicine, Beijing, P. R. China
| | - Yuji Ikeno
- Barshop Institute of Longevity and Aging Research, University of Texas Health San Antonio, San Antonio, Texas, USA
| | | | - Yidong Bai
- Barshop Institute of Longevity and Aging Research, University of Texas Health San Antonio, San Antonio, Texas, USA
- Department of Cell Systems and Anatomy, University of Texas Health San Antonio, 7703 Floyd Curl Drive, San Antonio, Texas, 78229, USA
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18
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Huang Q, Tang J, Chai X, Ren W, Wang J, Gan Q, Shi J, Wang M, Yang S, Liu J, Ma L. Affinity ultrafiltration and UPLC-HR-Orbitrap-MS based screening of thrombin-targeted small molecules with anticoagulation activity from Poecilobdella manillensis. J Chromatogr B Analyt Technol Biomed Life Sci 2021; 1178:122822. [PMID: 34147951 DOI: 10.1016/j.jchromb.2021.122822] [Citation(s) in RCA: 9] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/15/2020] [Revised: 05/06/2021] [Accepted: 05/31/2021] [Indexed: 01/19/2023]
Abstract
This study aims to screen potential anticoagulant components from leeches, a representative animal-sourced traditional Chinese medicine using thrombin (THR)-targeted ultrafiltration combined with ultrahigh performance liquid chromatography and high-resolution Orbitrap mass spectrometry (UPLC-HR-Orbitrap-MS). As a result, five small molecules in leech extract were discovered to interact with THR for the first time. Among them, two new compounds were isolated and their structures were identified by IR, HR-MS and NMR data. Furthermore, their THR inhibitory activity was confirmed with IC50 values of 4.74 and 8.31 μM, respectively. In addition, molecular docking analysis showed that the active (catalytic) site of THR might be the possible binding site of the two hits. Finally, reverse screening analysis indicated that LTA4-H, ACE and ALOX5AP were potential anticoagulant targets of the two new compounds. This study will broaden our understanding of the medicinal substance basis in leeches and further contribute to the discovery and development of clinical anticoagulant drugs from leeches.
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Affiliation(s)
- Qiuyang Huang
- School of Traditional Chinese Medicine, Capital Medical University, Beijing 100069, China
| | - Jianyuan Tang
- Hospital of Chengdu University of Traditional Chinese Medicine, Chengdu 610072, China
| | - Xiaoxin Chai
- School of Traditional Chinese Medicine, Capital Medical University, Beijing 100069, China
| | - Wei Ren
- Affiliated Traditional Chinese Medicine Hospital, Southwest Medical University, Luzhou 646000, China
| | - JiaBo Wang
- School of Traditional Chinese Medicine, Capital Medical University, Beijing 100069, China
| | - Qichao Gan
- Chongqing Duoputai Pharmaceutical Co., Ltd, Chongqing 400800, China
| | - Jingyan Shi
- Chongqing Duoputai Pharmaceutical Co., Ltd, Chongqing 400800, China
| | - Manyuan Wang
- School of Traditional Chinese Medicine, Capital Medical University, Beijing 100069, China
| | - Sijin Yang
- Affiliated Traditional Chinese Medicine Hospital, Southwest Medical University, Luzhou 646000, China
| | - Jingfang Liu
- Public Technology Service Center, Institute of Microbiology, Chinese Academy of Sciences, Beijing 100101, China
| | - Li Ma
- School of Traditional Chinese Medicine, Capital Medical University, Beijing 100069, China.
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19
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Muchiri RN, van Breemen RB. Affinity selection-mass spectrometry for the discovery of pharmacologically active compounds from combinatorial libraries and natural products. JOURNAL OF MASS SPECTROMETRY : JMS 2021; 56:e4647. [PMID: 32955158 DOI: 10.1002/jms.4647] [Citation(s) in RCA: 23] [Impact Index Per Article: 5.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Received: 05/04/2020] [Revised: 07/15/2020] [Accepted: 08/11/2020] [Indexed: 05/12/2023]
Abstract
Invented to address the high-throughput screening (HTS) demands of combinatorial chemistry, affinity selection-mass spectrometry (AS-MS) utilizes binding interactions between ligands and receptors to isolate pharmacologically active compounds from mixtures of small molecules and then relies on the selectivity, sensitivity, and speed of mass spectrometry to identify them. No radiolabels, fluorophores, or chromophores are required. Although many variations of AS-MS have been devised, three approaches have emerged as the most flexible, productive, and popular, and they differ primarily in how ligand-receptor complexes are separated from nonbinding compounds in the mixture. These are pulsed ultrafiltration (PUF) AS-MS, size exclusion chromatography (SEC) AS-MS, and magnetic microbead affinity selection screening (MagMASS). PUF and SEC AS-MS are solution-phase screening approaches, and MagMASS uses receptors immobilized on magnetic microbeads. Because pools of compounds are screened using AS-MS, each containing hundreds to thousands of potential ligands, hundreds of thousands of compounds can be screened per day. AS-MS is also compatible with complex mixtures of chemically diverse natural products in extracts of botanicals and fungi and microbial cultures, which often contain fluorophores and chromophores that can interfere with convention HTS. Unlike conventional HTS, AS-MS may be used to discover ligands binding to allosteric as well as orthosteric receptor sites, and AS-MS has been useful for discovering ligands to targets that are not easily incorporated into conventional HTS such as membrane-bound receptors.
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Affiliation(s)
- Ruth N Muchiri
- Linus Pauling Institute, Oregon State University, Corvallis, Oregon, 97331, USA
| | - Richard B van Breemen
- Linus Pauling Institute, Oregon State University, Corvallis, Oregon, 97331, USA
- Department of Pharmaceutical Sciences, Oregon State University, Corvallis, Oregon, 97331, USA
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20
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Tao Y, Yan J, Cai B. LABEL-FREE BIO-AFFINITY MASS SPECTROMETRY FOR SCREENING AND LOCATING BIOACTIVE MOLECULES. MASS SPECTROMETRY REVIEWS 2021; 40:53-71. [PMID: 31755145 DOI: 10.1002/mas.21613] [Citation(s) in RCA: 6] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Received: 04/17/2019] [Accepted: 11/04/2019] [Indexed: 06/10/2023]
Abstract
Despite the recent increase in the development of bioactive molecules in the drug industry, the enormous chemical space and lack of productivity are still important issues. Additional alternative approaches to screen and locate bioactive molecules are urgently needed. Label-free bio-affinity mass spectrometry (BA-MS) provides opportunities for the discovery and development of innovative drugs. This review provides a comprehensive portrayal of BA-MS techniques and of their applications in screening and locating bioactive molecules. After introducing the basic principles, alongside some application notes, the current state-of-the-art of BA-MS-assisted drug discovery is discussed, including native MS, size-exclusion chromatography-MS, ultrafiltration-MS, solid-phase micro-extraction-MS, and cell membrane chromatography-MS. Finally, several challenges and limitations of the current methods are summarized, with a view to potential future directions for BA-MS-assisted drug discovery. © 2019 John Wiley & Sons Ltd. Mass Spec Rev.
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Affiliation(s)
- Yi Tao
- College of Pharmaceutical Science, Zhejiang University of Technology, Hangzhou, 310032, PR China
| | - Jizhong Yan
- College of Pharmaceutical Science, Zhejiang University of Technology, Hangzhou, 310032, PR China
| | - Baochang Cai
- Jiangsu Key Laboratory of Chinese Medicine Processing, Nanjing University of Chinese Medicine, Nanjing, 210023, PR China
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21
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Halim ME, Omar MA, Abdallah NA, Nagy DM. The first spectrofluorimetric approach for accurate determination of trimetazidine in its dosage forms: Application to content uniformity testing. SPECTROCHIMICA ACTA. PART A, MOLECULAR AND BIOMOLECULAR SPECTROSCOPY 2020; 242:118710. [PMID: 32712572 DOI: 10.1016/j.saa.2020.118710] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Received: 05/25/2020] [Revised: 06/22/2020] [Accepted: 07/15/2020] [Indexed: 06/11/2023]
Abstract
A novel sensitive and simple spectrofluorimetric method has been developed then validated for the determination of trimetazidine in pure form and its tablets. This method is found on the reaction between trimetazidine's secondary amine moiety with NBD-Cl reagent, using borate buffer at pH 8.0 yielding a highly fluorescent product whose fluorescence intensity was measured at 526 nm (excitation at 466 nm). A calibration curve plotted showed that the linear range of the presented method was (50-700 ng/ml) with a correlation coefficient of 0.9998. The limits of detection (LOD) and limits of quantitation (LOQ) values were 15.01 and 45.50 ng/ml respectively. The presented approach was validated according to ICH guidelines and successfully applied for determining trimetazidine in its tablets with a mean percentage recovery of 99.65% ± 1.04, 99.23% ± 0.80 and 98.33% ± 1.03 for Metacardia® (20 mg), Vastor ® (20 mg) and Tricardia® (20 mg) tablets respectively. Finally, the proposed method was adopted to study the content uniformity test according to USP guidelines.
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Affiliation(s)
- Monica E Halim
- Analytical Chemistry Department, Faculty of Pharmacy, Minia University, Minia, Egypt
| | - Mahmoud A Omar
- Department of Pharmacognosy and Pharmaceutical Chemistry, College of Pharmacy, Taibah University, Medinah, Saudi Arabia; Pharmaceutical Analysis Department, Faculty of Pharmacy, Minia University, Minia, Egypt.
| | - Nehad A Abdallah
- Department of Pharmacognosy and Pharmaceutical Chemistry, College of Pharmacy, Taibah University, Medinah, Saudi Arabia
| | - Dalia M Nagy
- Analytical Chemistry Department, Faculty of Pharmacy, Minia University, Minia, Egypt
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Exploring new targets and chemical space with affinity selection-mass spectrometry. Nat Rev Chem 2020; 5:62-71. [PMID: 37118102 DOI: 10.1038/s41570-020-00229-2] [Citation(s) in RCA: 31] [Impact Index Per Article: 6.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Accepted: 09/16/2020] [Indexed: 12/15/2022]
Abstract
Affinity selection-mass spectrometry (AS-MS) is a high-throughput screening (HTS) technique for drug discovery that enables rapid screening of large collections of compounds to identify ligands for a specific biomolecular target. AS-MS is a binding assay that is insensitive to the functional effects a ligand might have, which is important because it lets us identify novel ligands irrespective of their binding site. This approach is gaining popularity, notably due to its role in the emergence of useful agents for targeted protein degradation. This Perspective highlights the use of AS-MS techniques to explore broad chemical space and identify small-molecule ligands for biological targets that have proven challenging to address with other screening paradigms. We present chemical structures of reported AS-MS hits to illustrate the potential of this screening approach to deliver high-quality hits for further optimization. AS-MS has, thus, evolved from being an infrequent alternative to traditional HTS or DNA-encoded library strategies to now firmly establishing itself as a HTS approach for drug discovery.
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Identification of Mitochondrial Ligands with Hepatoprotective Activity from Notopterygii Rhizoma et Radix Using Affinity Ultrafiltration/Liquid Chromatography/Mass Spectrometry. BIOMED RESEARCH INTERNATIONAL 2019; 2019:5729263. [PMID: 31950043 PMCID: PMC6948297 DOI: 10.1155/2019/5729263] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 07/02/2019] [Revised: 08/01/2019] [Accepted: 11/04/2019] [Indexed: 01/04/2023]
Abstract
In recent years, the incidence of diseases associated with hepatic injury has increased in prevalence. Targeting the mitochondria to protect liver function has gained momentum due to their central role in energy production, apoptotic cell death, oxidative stress, calcium homeostasis, and lipid metabolism. In this study, we employed a hepatic mitochondria-based centrifugal ultrafiltration/liquid chromatography/mass spectrometry method (CM-HMC) to identify hepatic mitochondria ligands from medicinal herbs (MHs) including Notopterygii Rhizoma et Radix (NRR) that possess hepatic-protective effects. A total of 4 newly identified mitochondrial ligands were successfully identified by CM-HMC. The mitochondria-regulating activities of 3 of the 4 hits were confirmed using isolated mitochondria. The hepatic-protective effects of one of these hits were validated in carbon tetrachloride-damaged human liver L02 cell models. We have thus identified new natural hepatic-protectants that enhance our understanding of the hepatic-protective mechanisms of MHs. CM-HMC was proven to efficiently screen for mitochondrial ligands from MHs.
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Yang XX, Wang X, Shi TT, Dong JC, Li FJ, Zeng LX, Yang M, Gu W, Li JP, Yu J. Mitochondrial dysfunction in high-fat diet-induced nonalcoholic fatty liver disease: The alleviating effect and its mechanism of Polygonatum kingianum. Biomed Pharmacother 2019; 117:109083. [PMID: 31387169 DOI: 10.1016/j.biopha.2019.109083] [Citation(s) in RCA: 31] [Impact Index Per Article: 5.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/16/2019] [Revised: 06/03/2019] [Accepted: 06/03/2019] [Indexed: 02/07/2023] Open
Abstract
BACKGROUND Mitochondrial dysfunction is an important mechanism of non-alcoholic fatty liver disease (NAFLD). Developing mitochondrial regulators/nutrients from natural products to remedy mitochondrial dysfunction represent attractive strategies for NAFLD therapy. In China, Polygonatum kingianum (PK) has been used as a herb and food nutrient for centuries. So far, studies in which the effects of PK on NAFLD are evaluated are lacking. Our study aims at identifying the effects and mechanism of action of PK on NAFLD based on mitochondrial regulation. METHODS A NAFLD rat model was induced by a high-fat diet (HFD) and rats were intragastrically given PK (1, 2 and 4 g/kg) for 14 weeks. Changes in body weight, food intake, histological parameters, organ indexes, biochemical parameters and mitochondrial indicators involved in oxidative stress, energy metabolism, fatty acid metabolism, and apoptosis were investigated. RESULTS PK significantly inhibited the HFD-induced increase of alanine transaminase, aspartate transaminase, total cholesterol (TC), and low density lipoprotein cholesterol in serum, and TC and triglyceride in the liver. In addition, PK reduced high density lipoprotein cholesterol and liver enlargement without affecting food intake. PK also remarkably inhibited the HFD-induced increase of malondialdehyde and the reduction of superoxide dismutase, glutathione peroxidase, ATP synthase, and complex I and II, in mitochondria. Moreover, mRNA expression of carnitine palmitoyl transferase-1 and uncoupling protein-2 was significantly up-regulated and down-regulated after PK treatment, respectively. Finally, PK notably inhibited the HFD-induced increase of caspase 9, caspase 3 and Bax expression in hepatocytes, and the decrease of expression of Bcl-2 in hepatocytes and cytchrome c in mitochondria. CONCLUSION PK alleviated HFD-induced NAFLD by promoting mitochondrial functions. Thus, PK may be useful mitochondrial regulators/nutrients to remedy mitochondrial dysfunction and alleviate NAFLD.
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Affiliation(s)
- Xing-Xin Yang
- College of Pharmaceutical Science, Yunnan University of Chinese Medicine, 1076 Yuhua Road, Kunming, 650500, China; Kunming Key Laboratory for Metabolic Diseases Prevention and Treatment by Chinese Medicine, 1076 Yuhua Road, Kunming, 650500, China
| | - Xi Wang
- College of Pharmaceutical Science, Yunnan University of Chinese Medicine, 1076 Yuhua Road, Kunming, 650500, China; Kunming Key Laboratory for Metabolic Diseases Prevention and Treatment by Chinese Medicine, 1076 Yuhua Road, Kunming, 650500, China
| | - Ting-Ting Shi
- Department of Pharmaceutical Preparation, The Xixi Hospital of Hangzhou Affiliated to Zhejiang University of Traditional Chinese medicine, Hangzhou, 310023, China
| | - Jin-Cai Dong
- College of Pharmaceutical Science, Yunnan University of Chinese Medicine, 1076 Yuhua Road, Kunming, 650500, China; Kunming Key Laboratory for Metabolic Diseases Prevention and Treatment by Chinese Medicine, 1076 Yuhua Road, Kunming, 650500, China
| | - Feng-Jiao Li
- College of Pharmaceutical Science, Yunnan University of Chinese Medicine, 1076 Yuhua Road, Kunming, 650500, China; Kunming Key Laboratory for Metabolic Diseases Prevention and Treatment by Chinese Medicine, 1076 Yuhua Road, Kunming, 650500, China
| | - Lin-Xi Zeng
- College of Pharmaceutical Science, Yunnan University of Chinese Medicine, 1076 Yuhua Road, Kunming, 650500, China; Kunming Key Laboratory for Metabolic Diseases Prevention and Treatment by Chinese Medicine, 1076 Yuhua Road, Kunming, 650500, China
| | - Min Yang
- College of Pharmaceutical Science, Yunnan University of Chinese Medicine, 1076 Yuhua Road, Kunming, 650500, China; Kunming Key Laboratory for Metabolic Diseases Prevention and Treatment by Chinese Medicine, 1076 Yuhua Road, Kunming, 650500, China
| | - Wen Gu
- College of Pharmaceutical Science, Yunnan University of Chinese Medicine, 1076 Yuhua Road, Kunming, 650500, China; Kunming Key Laboratory for Metabolic Diseases Prevention and Treatment by Chinese Medicine, 1076 Yuhua Road, Kunming, 650500, China
| | - Jing-Ping Li
- College of Pharmaceutical Science, Yunnan University of Chinese Medicine, 1076 Yuhua Road, Kunming, 650500, China; Kunming Key Laboratory for Metabolic Diseases Prevention and Treatment by Chinese Medicine, 1076 Yuhua Road, Kunming, 650500, China
| | - Jie Yu
- College of Pharmaceutical Science, Yunnan University of Chinese Medicine, 1076 Yuhua Road, Kunming, 650500, China; Kunming Key Laboratory for Metabolic Diseases Prevention and Treatment by Chinese Medicine, 1076 Yuhua Road, Kunming, 650500, China.
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Dexamethasone-Induced Mitochondrial Dysfunction and Insulin Resistance-Study in 3T3-L1 Adipocytes and Mitochondria Isolated from Mouse Liver. Molecules 2019; 24:molecules24101982. [PMID: 31126054 PMCID: PMC6572075 DOI: 10.3390/molecules24101982] [Citation(s) in RCA: 41] [Impact Index Per Article: 6.8] [Reference Citation Analysis] [Abstract] [Key Words] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/13/2019] [Revised: 05/14/2019] [Accepted: 05/21/2019] [Indexed: 02/01/2023] Open
Abstract
Dexamethasone is a glucocorticoid analog, which is reported to induce insulin resistance and to exacerbate diabetic symptoms. In this study, we investigated the association between mitochondrial dysfunction and the pathophysiology of dexamethasone-induced insulin resistance. An insulin resistance model in 3T3-L1 adipocyte was established by 48-h treatment of 1 μM dexamethasone, followed with the detection of mitochondrial function. Results showed that dexamethasone impaired insulin-induced glucose uptake and caused mitochondrial dysfunction. Abnormality in mitochondrial function was supported by decreased intracellular ATP and mitochondrial membrane potential (MMP), increased intracellular and mitochondrial reactive oxygen species (ROS) and mtDNA damage. Mitochondrial dynamic changes and biogenesis were suggested by decreased Drp1, increased Mfn2, and decreased PGC-1, NRF1, and TFam, respectively. The mitochondrial DNA (mtDNA) copy number exhibited no change while the mitochondrial mass increased. In agreement, studies in isolated mitochondria from mouse liver also showed dexamethasone-induced reduction of mitochondrial respiratory function, as suggested by decreased mitochondrial respiration controlling rate (RCR), lower MMP, declined ATP synthesis, opening of the mitochondrial permeability transition pore (mPTP), damage of mtDNA, and the accumulation of ROS. In summary, our study suggests that mitochondrial dysfunction occurs along with dexamethasone-induced insulin resistance in 3T3 L1 adipocytes and might be a potential mechanism of dexamethasone-induced insulin resistance.
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Wang HQ, Zhu YX, Liu YN, Wang RL, Wang SF. Rapid discovery and identification of the anti-inflammatory constituents in Zhi-Shi-Zhi-Zi-Chi-Tang. Chin J Nat Med 2019; 17:308-320. [PMID: 31076135 DOI: 10.1016/s1875-5364(19)30035-4] [Citation(s) in RCA: 6] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/12/2018] [Indexed: 01/01/2023]
Abstract
The anti-inflammatory active ingredients of Zhi-Shi-Zhi-Zi-Chi-Tang (ZSZZCT), a traditional Chinese medicine formula, were predicted and identified using an approach based on activity index, LC-MS, semi-preparative LC and NMR. Firstly, the whole extract of ZSZZCT was analyzed using liquid chromatography-quadrupole time of flight-mass spectrometry (LC-Q-TOF-MS) and liquid chromatography - ion trap mass spectrometry (LC-IT-MS), 79 constituents were detected and 39 constituents were identified unambiguously or tentatively. Subsequently, the whole extract of the formula was separated into multiple components and the activity index method was used to calculate index values of the 79 constituents by integrating the chemical and pharmacological information of multiple components. Four polymethoxyl flavones were predicted as the major active constituents according to the activity index values. Furthermore, three polymethoxyl flavones were prepared using the strategy with semi-preparative LC guided by LC-MS, and their anti-inflammatory activities were validated. The results show that three polymethoxyl flavones with higher positive index values, i.e., 3, 5, 6, 7, 8, 3', 4'-heptamethoxyflavone, 3-hydroxynobiletein and tangeretin had significant anti-inflammatory effects. In conclusion, the predicted results indicated that the activity index method is feasible for the accurate prediction of active constituents in TCM formulae.
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Affiliation(s)
- Hai-Qiang Wang
- College of Pharmaceutical Sciences, Zhejiang University, Hangzhou 310063, China
| | - Yun-Xiang Zhu
- College of Pharmaceutical Sciences, Zhejiang University, Hangzhou 310063, China
| | - Yi-Ning Liu
- College of Pharmaceutical Sciences, Zhejiang University, Hangzhou 310063, China
| | - Ruo-Liu Wang
- College of Pharmaceutical Sciences, Zhejiang University, Hangzhou 310063, China
| | - Shu-Fang Wang
- College of Pharmaceutical Sciences, Zhejiang University, Hangzhou 310063, China.
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Efficiently Capturing Mitochondria-Targeted Constituents with Hepatoprotective Activity from Medicinal Herbs. OXIDATIVE MEDICINE AND CELLULAR LONGEVITY 2019; 2019:4353791. [PMID: 31093314 PMCID: PMC6481013 DOI: 10.1155/2019/4353791] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 09/29/2018] [Revised: 12/20/2018] [Accepted: 01/16/2019] [Indexed: 12/13/2022]
Abstract
Targeting mitochondria as a hepatic-protective strategy has gained attention, because of their important roles in energy production, adjustment of apoptosis, and generation of reactive oxygen species. To promote the discovery of natural mitochondria-targeted hepatic-protectants, we established a hepatocellular mitochondria-based capturing method by coupling affinity ultrafiltration with liquid chromatography/mass spectrometry (LC/MS), which is suitable for identifying mitochondrial ligands from medicinal herbs (MHs). After evaluating the feasibility of the method, it was applied for capturing mitochondria-targeting constituents from Peucedani Radix extract. A total of 10 active compounds were identified by LC/MS, all of which were newly identified mitochondrial ligands. The mitochondria-remedying activity of 4 of the 10 hits was confirmed by pharmacological tests in vitro. Additionally, the hepatic-protective abilities of 4 hits were verified in both carbon tetrachloride-damaged liver L02 cells and mice. These results indicated that the method could be used for identifying hepatic mitochondria-targeting constituents in MHs, which might be beneficial for hepatic-protective development.
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Zhang J, Yang C, Pan S, Shi M, Li J, Hu H, Qiao M, Chen D, Zhao X. Eph A10-modified pH-sensitive liposomes loaded with novel triphenylphosphine-docetaxel conjugate possess hierarchical targetability and sufficient antitumor effect both in vitro and in vivo. Drug Deliv 2018. [PMID: 29513049 PMCID: PMC6058733 DOI: 10.1080/10717544.2018.1446475] [Citation(s) in RCA: 24] [Impact Index Per Article: 3.4] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/25/2022] Open
Abstract
Mitochondrial-targeting therapy was considered to be a promising approach for the efficient treatment of cancer while positive charge induced nonspecific cytotoxicity severely limits its application. To overcome this drawback, a novel mitochondria targeted conjugate triphenylphosphine-docetaxel (TD) has been synthesized successfully and incorporated it into liposomes (EPSLP/TD), which possessed excellent pH-sensitive characteristic, EphA 10 mediated active targetability as well as mitochondria-targeting capability. EPSLP/TD was characterized to have a small particle size, high-encapsulation efficiency and excellent pH-sensitive characteristic. Compared with DTX-loaded liposomes (EPSLP/DTX), EPSLP/TD possessed higher cytotoxicity against MCF-7 cell line. Mitochondrial-targeting assay demonstrated mitochondria-targeting moiety triphenylphosphine (TPP) could efficiently deliver DTX to mitochondria. Western immunoblotting assay indicated that EPSLP/TD could efficiently deliver antitumor drug to mitochondria and induce cell apoptosis via mitochondria-mediated apoptosis pathway. In vivo antitumor study demonstrated EPSLP/TD owed excellent in vivo antitumor activity. Histological assay demonstrated EPSLP/TD showed strongly apoptosis inducing effect, anti-proliferation effect and anti-angiogenesis effect. This work investigated the potential of hierarchical targeting pH-sensitive liposomes is a suitable carrier to activate mitochondria-mediated apoptosis pathway for cancer therapy.
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Affiliation(s)
- Jiulong Zhang
- a School of Pharmacy , Shenyang Pharmaceutical University , Shenyang , Liaoning , PR China
| | - Chunrong Yang
- b College Pharmacy , Jiamusi University , Jiamusi , Heilongjiang , PR China
| | - Shuang Pan
- a School of Pharmacy , Shenyang Pharmaceutical University , Shenyang , Liaoning , PR China
| | - Menghao Shi
- a School of Pharmacy , Shenyang Pharmaceutical University , Shenyang , Liaoning , PR China
| | - Jie Li
- c Mudanjiang Medical University , Mudanjiang , Heilongjiang , PR China
| | - Haiyang Hu
- a School of Pharmacy , Shenyang Pharmaceutical University , Shenyang , Liaoning , PR China
| | - Mingxi Qiao
- a School of Pharmacy , Shenyang Pharmaceutical University , Shenyang , Liaoning , PR China
| | - Dawei Chen
- a School of Pharmacy , Shenyang Pharmaceutical University , Shenyang , Liaoning , PR China
| | - Xiuli Zhao
- a School of Pharmacy , Shenyang Pharmaceutical University , Shenyang , Liaoning , PR China
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Chen G, Huang BX, Guo M. Current advances in screening for bioactive components from medicinal plants by affinity ultrafiltration mass spectrometry. PHYTOCHEMICAL ANALYSIS : PCA 2018; 29:375-386. [PMID: 29785715 DOI: 10.1002/pca.2769] [Citation(s) in RCA: 41] [Impact Index Per Article: 5.9] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Received: 12/27/2017] [Revised: 03/08/2018] [Accepted: 03/08/2018] [Indexed: 06/08/2023]
Abstract
INTRODUCTION Medicinal plants have played an important role in maintaining human health for thousands of years. However, the interactions between the active components in medicinal plants and some certain biological targets during a disease are still unclear in most cases. OBJECTIVE To conduct the high-throughput screening for small active molecules that can interact with biological targets, which is of great theoretical significance and practical value. METHODOLOGY The ultrafiltration mass spectrometry (UF-LC/MS) is a powerful bio-analytical method by combining affinity ultrafiltration and liquid chromatography-mass spectrometry (LC/MS), which could rapidly screen and identify small active molecules that bind to biological targets of interest at the same time. Compared with other analytical methods, affinity UF-LC/MS has the characteristics of fast, sensitive and high throughput, and is especially suitable for the complicated extracts of medicinal plants. RESULTS In this review, the basic principle, characteristics and some most recent challenges in UF-LC/MS have been demonstrated. Meanwhile, the progress and applications of affinity UF-LC/MS in the discovery of the active components from natural medicinal plants and the interactions between small molecules and biological target proteins are also briefly summarised. In addition, the future directions for UF-LC/MS are also prospected. CONCLUSION Affinity UF-LC/MS is a powerful tool in studies on the interactions between small active molecules and biological protein targets, especially in the high-throughput screening of active components from the natural medicinal plants.
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Affiliation(s)
- Guilin Chen
- Key Laboratory of Plant Germplasm Enhancement and Specialty Agriculture, Wuhan Botanical Garden, Chinese Academy of Sciences, 430074, Wuhan, China
- Sino-Africa Joint Research Center, Chinese Academy of Sciences, 430074, Wuhan, China
| | - Bill X Huang
- Laboratory of Molecular Signaling, National Institute of Alcohol Abuse and Alcoholism, National Institutes of Health, Bethesda, Maryland, USA
| | - Mingquan Guo
- Key Laboratory of Plant Germplasm Enhancement and Specialty Agriculture, Wuhan Botanical Garden, Chinese Academy of Sciences, 430074, Wuhan, China
- Sino-Africa Joint Research Center, Chinese Academy of Sciences, 430074, Wuhan, China
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Yang XX, Zhou YZ, Xu F, Yu J, Gegentana, Shang MY, Wang X, Cai SQ. Screening potential mitochondria-targeting compounds from traditional Chinese medicines using a mitochondria-based centrifugal ultrafiltration/liquid chromatography/mass spectrometry method. J Pharm Anal 2018; 8:240-249. [PMID: 30140488 PMCID: PMC6104153 DOI: 10.1016/j.jpha.2018.06.001] [Citation(s) in RCA: 11] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/08/2017] [Revised: 06/07/2018] [Accepted: 06/08/2018] [Indexed: 12/16/2022] Open
Abstract
Mitochondria regulate numerous crucial cell processes, including energy production, apoptotic cell death, oxidative stress, calcium homeostasis and lipid metabolism. Here, we applied an efficient mitochondria-based centrifugal ultrafiltration/liquid chromatography/mass spectrometry (LC/MS) method, also known as screening method for mitochondria-targeted bioactive constituents (SM-MBC). This method allowed searching natural mitochondria-targeting compounds from traditional Chinese medicines (TCMs), including Puerariae Radix (PR) and Chuanxiong Radix (CR). A total of 23 active compounds were successfully discovered from the two TCMs extracts. Among these 23 hit compounds, 17 were identified by LC/MS, 12 of which were novel mitochondria-targeting compounds. Among these, 6 active compounds were analyzed in vitro for pharmacological tests and found able to affect mitochondrial functions. We also investigated the effects of the hit compounds on HepG2 cell proliferation and on loss of cardiomyocyte viability induced by hypoxia/reoxygenation injury. The results obtained are useful for in-depth understanding of mechanisms underlying TCMs therapeutic effects at mitochondria level and for developing novel potential drugs using TCMs as lead compounds. Finally, we showed that SM-MBC was an efficient protocol for the rapid screening of mitochondria-targeting constituents from complex samples such as PR and CR extracts.
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Affiliation(s)
- Xing-Xin Yang
- Department of Natural Medicines, School of Pharmaceutical Sciences, Peking University, 38 Xueyuan Road, Beijing 100191, PR China.,Department of Chemical Biology, School of Pharmaceutical Sciences, Peking University, 38 Xueyuan Road, Beijing 100191, PR China.,College of Pharmaceutical Science, Yunnan University of Traditional Chinese Medicine, 1076 Yuhua Road, Kunming 650500, Yunnan Province, PR China
| | - Yu-Zhen Zhou
- Department of Chemical Biology, School of Pharmaceutical Sciences, Peking University, 38 Xueyuan Road, Beijing 100191, PR China
| | - Feng Xu
- Department of Natural Medicines, School of Pharmaceutical Sciences, Peking University, 38 Xueyuan Road, Beijing 100191, PR China
| | - Jie Yu
- College of Pharmaceutical Science, Yunnan University of Traditional Chinese Medicine, 1076 Yuhua Road, Kunming 650500, Yunnan Province, PR China
| | - Gegentana
- Department of Natural Medicines, School of Pharmaceutical Sciences, Peking University, 38 Xueyuan Road, Beijing 100191, PR China
| | - Ming-Ying Shang
- Department of Natural Medicines, School of Pharmaceutical Sciences, Peking University, 38 Xueyuan Road, Beijing 100191, PR China
| | - Xuan Wang
- Department of Chemical Biology, School of Pharmaceutical Sciences, Peking University, 38 Xueyuan Road, Beijing 100191, PR China
| | - Shao-Qing Cai
- Department of Natural Medicines, School of Pharmaceutical Sciences, Peking University, 38 Xueyuan Road, Beijing 100191, PR China
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Wang YL, Hu G, Zhang Q, Yang YX, Li QQ, Hu YJ, Chen H, Yang FQ. Screening and Characterizing Tyrosinase Inhibitors from Salvia miltiorrhiza and Carthamus tinctorius by Spectrum-Effect Relationship Analysis and Molecular Docking. JOURNAL OF ANALYTICAL METHODS IN CHEMISTRY 2018; 2018:2141389. [PMID: 29862119 PMCID: PMC5971358 DOI: 10.1155/2018/2141389] [Citation(s) in RCA: 10] [Impact Index Per Article: 1.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Subscribe] [Scholar Register] [Received: 02/13/2018] [Accepted: 04/01/2018] [Indexed: 05/08/2023]
Abstract
Tyrosinase (TYR) is a rate-limiting enzyme in the synthesis of melanin, while direct TYR inhibitors are a class of important clinical antimelanoma drugs. This study established a spectrum-effect relationship analysis method and high-performance liquid chromatography-mass spectrometry (LC-MS) analysis method to screen and identify the active ingredients that inhibited TYR in Salvia miltiorrhiza-Carthamus tinctorius (Danshen-Honghua, DH) herbal pair. Seventeen potential active compounds (peaks) in the extract of DH herbal pair were predicted, and thirteen of them were tentatively identified by LC-MS analysis. Furthermore, TYR inhibitory activities of five pure compounds obtained from the DH herbal pair were validated in the test in which kojic acid served as a positive control drug. Among them, three compounds including protocatechuic aldehyde, hydroxysafflor yellow A, and tanshinone IIA were verified to have high TYR inhibitory activity (IC50 value of 455, 498, and 1214 μM, resp.) and bind to the same amino acid residues in TYR catalytic pocket according to the results of the molecular docking test. However, the other two compounds lithospermic acid and salvianolic acid A had a weak effect on TYR, as they do not combine with the active amino acid residues or act on the active center of TYR. Therefore, the developed methods (spectrum-effect relationship analysis and molecular docking) could be used to effectively screen TYR inhibitors in complex mixtures such as natural products.
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Affiliation(s)
- Ya-Li Wang
- School of Pharmacy and Bioengineering, Chongqing University of Technology, Chongqing 400054, China
- School of Chemistry and Chemical Engineering, Chongqing University, Chongqing 401331, China
| | - Guang Hu
- School of Pharmacy and Bioengineering, Chongqing University of Technology, Chongqing 400054, China
| | - Qian Zhang
- School of Chemistry and Chemical Engineering, Chongqing University, Chongqing 401331, China
| | - Yu-Xiu Yang
- School of Chemistry and Chemical Engineering, Chongqing University, Chongqing 401331, China
| | - Qiao-Qiao Li
- School of Chemistry and Chemical Engineering, Chongqing University, Chongqing 401331, China
| | - Yuan-Jia Hu
- State Key Laboratory of Quality Research in Chinese Medicine, Institute of Chinese Medical Sciences, University of Macau, Macau
| | - Hua Chen
- School of Chemistry and Chemical Engineering, Chongqing University, Chongqing 401331, China
| | - Feng-Qing Yang
- School of Chemistry and Chemical Engineering, Chongqing University, Chongqing 401331, China
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Recent developments and emerging trends of mass spectrometry for herbal ingredients analysis. Trends Analyt Chem 2017. [DOI: 10.1016/j.trac.2017.07.007] [Citation(s) in RCA: 60] [Impact Index Per Article: 7.5] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/22/2022]
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An ultrafiltration and high performance liquid chromatography coupled with diode array detector and mass spectrometry approach for screening and characterizing thrombin inhibitors from Rhizoma Chuanxiong. J Chromatogr B Analyt Technol Biomed Life Sci 2017; 1061-1062:421-429. [PMID: 28818799 DOI: 10.1016/j.jchromb.2017.07.050] [Citation(s) in RCA: 29] [Impact Index Per Article: 3.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/07/2017] [Revised: 07/24/2017] [Accepted: 07/28/2017] [Indexed: 12/11/2022]
Abstract
Thrombin (THR) plays a significant role in thromboembolic diseases, direct THR inhibitors are a class of important clinical anticoagulant drugs. This study established a THR in-solution based biospecific extraction combined with ultrafiltration and high performance liquid chromatography coupled with diode array detector and mass spectrometry analysis (TUA) method to screen and identify ligands for THR in Rhizoma Chuanxiong. After evaluating the reliability of the present TUA method using positive (argatroban) and negative (adenosine, tirofiban, ticagrelor) control drugs, this method was successfully applied to detect eight potential active compounds in Rhizoma Chuanxiong. Two new THR-targeted compounds isochlorogenic acid C and senkyunolide I with high THR inhibitory activity (IC50 206.48 and 197.23μM, respectively) were identified by liquid chromatography/mass spectrometry and enzyme inhibitory activity test finally. They were reported with direct THR inhibition activity for the first time and their ligand-THR interactions were explored by in silico molecular docking research. In addition, based on the TUA screening result, four compounds gained similar structure with the two hit compounds were also investigated as promising candidates targeting THR with high binding energy (>5.0kcal/mol). These results may prove that the proposed method could effectively screen THR inhibitors in complex mixtures.
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A novel pretreatment method combining sealing technique with direct injection technique applied for improving biosafety. Bioanalysis 2017; 9:173-182. [PMID: 27960549 DOI: 10.4155/bio-2016-0154] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/17/2022] Open
Abstract
AIM People today have a stronger interest in the risk of biosafety in clinical bioanalysis. A safe, simple, effective method of preparation is needed urgently. METHODOLOGY/RESULTS To improve biosafety of clinical analysis, we used antiviral drugs of adefovir and tenofovir as model drugs and developed a safe pretreatment method combining sealing technique with direct injection technique. The inter- and intraday precision (RSD %) of the method were <4%, and the extraction recoveries ranged from 99.4 to 100.7%. Meanwhile, the results showed that standard solution could be used to prepare calibration curve instead of spiking plasma, acquiring more accuracy result. CONCLUSION/DISCUSSION Compared with traditional methods, the novel method not only improved biosecurity of the pretreatment method significantly, but also achieved several advantages including higher precision, favorable sensitivity and satisfactory recovery. With these highly practical and desirable characteristics, the novel method may become a feasible platform in bioanalysis.
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Yang XX, Gu W, Liang L, Yan HL, Wang YF, Bi Q, Zhang T, Yu J, Rao GX. Screening for the bioactive constituents of traditional Chinese medicines—progress and challenges. RSC Adv 2017. [DOI: 10.1039/c6ra25765h] [Citation(s) in RCA: 10] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/02/2023] Open
Abstract
The search for lead compounds from traditional Chinese medicines (TCMs) may be promising for new drug development.
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Affiliation(s)
- Xing-Xin Yang
- College of Pharmaceutical Science
- Yunnan University of Traditional Chinese Medicine
- Kunming 650500
- P. R. China
| | - Wen Gu
- College of Pharmaceutical Science
- Yunnan University of Traditional Chinese Medicine
- Kunming 650500
- P. R. China
| | - Li Liang
- College of Pharmaceutical Science
- Yunnan University of Traditional Chinese Medicine
- Kunming 650500
- P. R. China
| | - Hong-Li Yan
- College of Pharmaceutical Science
- Yunnan University of Traditional Chinese Medicine
- Kunming 650500
- P. R. China
| | - Yan-Fang Wang
- College of Pharmaceutical Science
- Yunnan University of Traditional Chinese Medicine
- Kunming 650500
- P. R. China
| | - Qian Bi
- College of Pharmaceutical Science
- Yunnan University of Traditional Chinese Medicine
- Kunming 650500
- P. R. China
| | - Ting Zhang
- College of Pharmaceutical Science
- Yunnan University of Traditional Chinese Medicine
- Kunming 650500
- P. R. China
| | - Jie Yu
- College of Pharmaceutical Science
- Yunnan University of Traditional Chinese Medicine
- Kunming 650500
- P. R. China
- Engineering Laboratory for National Healthcare Theories and Products of Yunnan Province
| | - Gao-Xiong Rao
- College of Pharmaceutical Science
- Yunnan University of Traditional Chinese Medicine
- Kunming 650500
- P. R. China
- Engineering Laboratory for National Healthcare Theories and Products of Yunnan Province
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Cieśla Ł, Moaddel R. Comparison of analytical techniques for the identification of bioactive compounds from natural products. Nat Prod Rep 2016; 33:1131-45. [PMID: 27367973 PMCID: PMC5042860 DOI: 10.1039/c6np00016a] [Citation(s) in RCA: 89] [Impact Index Per Article: 9.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/22/2022]
Abstract
Covering: 2000 to 2016Natural product extracts are a rich source of bioactive compounds. As a result, the screening of natural products for the identification of novel biologically active metabolites has been an essential part of several drug discovery programs. It is estimated that more than 70% of all drugs approved from 1981 and 2006, were either derived from or structurally similar to nature based compounds indicating the necessity for the development of a rapid method for the identification of novel compounds from plant extracts. The screening of biological matrices for the identification of novel modulators is nevertheless still challenging. In this review we discuss current techniques in phytochemical analysis and the identification of biologically active components.
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Affiliation(s)
- Łukasz Cieśla
- Laboratory of Clinical Investigation, Biomedical Research Center, 8C232, National Institute on Aging, National Institutes of Health, 251 Bayview Boulevard, Baltimore, Maryland 21224, USA.
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Wei H, Zhang X, Tian X, Wu G. Pharmaceutical applications of affinity-ultrafiltration mass spectrometry: Recent advances and future prospects. J Pharm Biomed Anal 2016; 131:444-453. [PMID: 27668554 DOI: 10.1016/j.jpba.2016.09.021] [Citation(s) in RCA: 46] [Impact Index Per Article: 5.1] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/30/2016] [Revised: 09/06/2016] [Accepted: 09/20/2016] [Indexed: 11/17/2022]
Abstract
The immunoaffinity of protein with ligand is broadly involved in many bioanalytical methods. Affinity-ultrafiltration mass spectrometry (AUF-MS), a platform based on interaction of protein-ligand affinity, has been developed to fish out interesting molecules from complex matrixes. Here we reviewed the basics of AUF-MS and its recent applications to pharmaceutical field, i.e. target-oriented discovery of lead compounds from combinatorial libraries and natural product extracts, and determination of free drug concentration in biosamples. Selected practical examples were highlighted to illustrate the advances of AUF-MS in pharmaceutical fields. The future prospects were also presented.
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Affiliation(s)
- Han Wei
- Department of Pharmacy, The First Affiliated Hospital of Zhengzhou University, Zhengzhou 450052, China.
| | - Xiaojian Zhang
- Department of Pharmacy, The First Affiliated Hospital of Zhengzhou University, Zhengzhou 450052, China
| | - Xin Tian
- Department of Pharmacy, The First Affiliated Hospital of Zhengzhou University, Zhengzhou 450052, China
| | - Guanghua Wu
- Department of Pharmacy, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030, China.
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Yu H, Lu JJ, Rao W, Liu S. Capitalizing Resolving Power of Density Gradient Ultracentrifugation by Freezing and Precisely Slicing Centrifuged Solution: Enabling Identification of Complex Proteins from Mitochondria by Matrix Assisted Laser Desorption/Ionization Time-of-Flight Mass Spectrometry. JOURNAL OF ANALYTICAL METHODS IN CHEMISTRY 2016; 2016:8183656. [PMID: 27668122 PMCID: PMC5030451 DOI: 10.1155/2016/8183656] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [Abstract] [Grants] [Track Full Text] [Figures] [Subscribe] [Scholar Register] [Received: 07/13/2016] [Revised: 08/01/2016] [Accepted: 08/07/2016] [Indexed: 06/06/2023]
Abstract
Density gradient centrifugation is widely utilized for various high purity sample preparations, and density gradient ultracentrifugation (DGU) is often used for more resolution-demanding purification of organelles and protein complexes. Accurately locating different isopycnic layers and precisely extracting solutions from these layers play a critical role in achieving high-resolution DGU separations. In this technique note, we develop a DGU procedure by freezing the solution rapidly (but gently) after centrifugation to fix the resolved layers and by slicing the frozen solution to fractionate the sample. Because the thickness of each slice can be controlled to be as thin as 10 micrometers, we retain virtually all the resolution produced by DGU. To demonstrate the effectiveness of this method, we fractionate complex V from HeLa mitochondria using a conventional technique and this freezing-slicing (F-S) method. The comparison indicates that our F-S method can reduce complex V layer thicknesses by ~40%. After fractionation, we analyze complex V proteins directly on a matrix assisted laser desorption/ionization, time-of-flight mass spectrometer. Twelve out of fifteen subunits of complex V are positively identified. Our method provides a practical protocol to identify proteins from complexes, which is useful to investigate biomolecular complexes and pathways in various conditions and cell types.
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Affiliation(s)
- Haiqing Yu
- Department of Chemistry and Biochemistry, University of Oklahoma, 101 Stephenson Parkway, Norman, OK 73019, USA
| | - Joann J. Lu
- Department of Chemistry and Biochemistry, University of Oklahoma, 101 Stephenson Parkway, Norman, OK 73019, USA
| | - Wei Rao
- Department of Chemistry and Biochemistry, University of Oklahoma, 101 Stephenson Parkway, Norman, OK 73019, USA
| | - Shaorong Liu
- Department of Chemistry and Biochemistry, University of Oklahoma, 101 Stephenson Parkway, Norman, OK 73019, USA
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Analysis of Bovine Serum Albumin Ligands from Puerariae flosUsing Ultrafiltration Combined with HPLC-MS. J CHEM-NY 2015. [DOI: 10.1155/2015/648361] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/17/2022] Open
Abstract
Rapid screening techniques for identification of active compounds from natural products are important not only for clarification of the therapeutic material basis, but also for supplying suitable chemical markers for quality control. In the present study, ultrafiltration combined with high performance liquid chromatography-mass spectrometry (HPLC-MS) was developed and conducted to screen and identify bovine serum albumin (BSA) bound ligands fromPuerariae flos. Fundamental parameters affecting the screening like incubation time, BSA concentration, pH, and temperature were studied and optimized. Under the optimum conditions, nine active compounds were identified by UV and MS data. The results indicated that this method was able to screen and identify BSA bound ligands form natural products without the need of preparative isolation techniques. Moreover, the method has more effective with easier operation procedures.
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