Size exclusion chromatographic analysis of polyphenol–serum albumin complexes

Hatano, Tsutomu; Hori, Mami; Hemingway, Richard W.; Yoshida, Takashi

doi:10.1016/s0031-9422(03)00340-6

Cited by 39 publications

(29 citation statements)

References 22 publications

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“…For example, EGCG has been reported to form water-soluble complexes with bovine serum albumin. 17) We showed that the anti-HCV activity of EGCG is diminished in the presence of serum. Thus, serumfree medium offers various benefits to screening programs.…”

Section: Discussionmentioning

confidence: 86%

A Cell-Based, Microplate Colorimetric Screen Identifies 7,8-Benzoflavone and Green Tea Gallate Catechins as Inhibitors of the Hepatitis C Virus

Fukazawa

Suzuki

Wakita

et al. 2012

Biological & Pharmaceutical Bulletin

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We describe a cell-based, microplate colorimetric screen for anti-hepatitis C virus (HCV) drugs that exploits the HCV-JFH1 viral culture system. Antiviral activity was assessed by measuring protection against the HCV-JFH1-induced cytopathic effect (CPE) in Huh7.5.1 cells using the 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide (MTT) viability assay. The use of serum-free medium substantially sensitized Huh7.5.1 cells to HCV-induced CPE, causing sufficient cell death to perform colorimetric assays for anti-HCV activity in 96-well plates. As a proof of concept, we carried out a pilot screen of an inhibitor library and identified cyclosporin A and tamoxifen, two compounds with reported anti-HCV activity. Using the assay, we discovered the anti-HCV properties of the plant flavonoids epigallocatechin gallate (EGCG) and 7,8-benzoflavone (α-naphthoflavone). Other gallate-type catechins and flavones also displayed anti-HCV activity, but 5,6-benzoflavone (β-naphthoflavone), flavanone, and non-gallate catechins were inactive. EGCG apparently acted mainly on HCV entry, although it may also block other steps. In contrast, 7,8-benzoflavone was presumed to inhibit later stages of the HCV life cycle. This assay is simple, reliable and cost-effective; does not require any specially engineered cell lines or viruses; and should be useful in the identification of compounds with anti-HCV activity. Key words hepatitis C virus; 7,8-benzoflavone (α-naphthoflavone); epigallocatechin gallateMore than 170 million people worldwide are chronically infected with the hepatitis C virus (HCV) and are at risk for developing liver diseases such as cirrhosis and hepatocellular carcinoma. Vaccines against HCV are not currently available; furthermore, the standard interferon/ribavirin combination therapy is not effective in approximately half of HCV-infected patients, and it has considerable side effects.1,2) Thus, there is an obvious and urgent need for new agents that can enhance or replace current HCV therapies.Screening programs using HCV replicon-based systems have successfully identified compounds that act on HCV RNA replication. However, replicon systems do not reproduce the entire HCV life cycle, and they cannot isolate inhibitors of many important steps such as viral entry, assembly, and egress. HCV cell culture infection models that recapitulate the entire viral life cycle in vitro have greatly enhanced the opportunity for HCV drug discovery. Several reports, including ours, have demonstrated that this model can overcome the limitations of the HCV replicon system and enable the discovery of compounds that target various stages of the HCV life cycle. [3][4][5][6][7][8] Using the HCV cell culture system, we previously developed a tube-capture-reverse transcription-polymerase chain reaction (RT-PCR) assay for screening HCV inhibitors and identified bisindolylmaleimides and indolocarbazoles as inhibitors of HCV replication.4) Here, we describe another screening method for the detection of anti-HCV activity. This assay measures t...

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Section: Discussionmentioning

confidence: 86%

A Cell-Based, Microplate Colorimetric Screen Identifies 7,8-Benzoflavone and Green Tea Gallate Catechins as Inhibitors of the Hepatitis C Virus

Fukazawa

Suzuki

Wakita

et al. 2012

Biological & Pharmaceutical Bulletin

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“…Binding affinity of pyrogallol-type catechins with HSA in aqueous solutions Several analytical approaches, including electrophoresis, spectrofluorometry, and chromatography, have indicated that catechins noncovalently bind to serum albumin. [22][23][24] HPLC with an HSA column recently enabled us to show that the hydroxyl groups on the galloyl group of each catechin influenced the binding affinity for immobilized HSA in an acidic solution with 20% acetonitrile. 25) We investigated in a QCM in the present study the interaction between pyrogallol-type catechins (EGC and EGCg) and HSA immobilized under physiological conditions.…”

Section: Resultsmentioning

confidence: 99%

Human Serum Albumin as an Antioxidant in the Oxidation of (−)-Epigallocatechin Gallate: Participation of Reversible Covalent Binding for Interaction and Stabilization

Ishii

Ichikawa

Minoda

et al. 2011

Bioscience, Biotechnology, and Biochemistry

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“…As a good chelator, EGCG Pk# may chelate with amino acid, protein, metal ion and so on. Hatano et al(2003)'s study showed that albumin of fetal bovine serum chelated with EGCG in cell culture system. Probably, chelation of EGCG may be the main reaction existing in the system.…”

Section: Change Of Egcg Content In F-12 Mediummentioning

confidence: 99%

Cytotoxicity of epigallocatechin-3-gallate to LNCaP cells in the presence of Cu2+

Shen

Xiong³

2005

J Zheijang Univ Sci B

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Epigallocatechin-3-gallate (EGCG) has shown remarkably anti-cancer activity, with its bioactivity being related to reactive conditions, such as pH and metal ions. The present study investigated the degradation of EGCG and its effect on prostate cancer cell in the presence of Cu 2+. EGCG was incubated with prostate cancer cells, LNCaP, pretreated with or without Cu 2+ . EGCG in F-12 medium was quantified using HPLC and the viability of cells was assessed by gel electrophoresis, flow cytometry, and electron microscope. The results of HPLC showed that EGCG degraded completely within 12 h in F-12 medium with or without Cu 2+. Gel electrophoresis and flow cytometry did not detect apoptosis of LNCaP cells when they were incubated with EGCG. Electron microscopy examination revealed that EGCG-Cu 2+ complex led to damage of cytoplasm membrane in LNCaP cells. It was speculated that not EGCG, but its oxide and complex with Cu 2+ , are the bioactive components responsible for its cytotoxicity to LNCaP prostate cancer cells.

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Size exclusion chromatographic analysis of polyphenol–serum albumin complexes

Cited by 39 publications

References 22 publications

A Cell-Based, Microplate Colorimetric Screen Identifies 7,8-Benzoflavone and Green Tea Gallate Catechins as Inhibitors of the Hepatitis C Virus

A Cell-Based, Microplate Colorimetric Screen Identifies 7,8-Benzoflavone and Green Tea Gallate Catechins as Inhibitors of the Hepatitis C Virus

Human Serum Albumin as an Antioxidant in the Oxidation of (−)-Epigallocatechin Gallate: Participation of Reversible Covalent Binding for Interaction and Stabilization

Cytotoxicity of epigallocatechin-3-gallate to LNCaP cells in the presence of Cu2+

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