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World Health Organization, Communicable Disease Surveillance & Response, website: http://www.who.int/csr/sars/archive/2003_05_07a/en and http://www.who.int/csr/sars/country/en/country2003_08_15.pdf.
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14
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53349162341
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note
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Thus far, no antiviral activity has been reported for these active ester-derived inhibitors. Inhibitor 3 did not exhibit any antiviral activity in our assay.
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15
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12144276705
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The indolecarboxylic acids are either from a commercial source or made from hydrolysis of corresponding esters or oxidation of corresponding aldehydes. The starting material for 13 is from Boc protection of 6-aminopenicillanic acid. For preparation of the starting material of 14, 2-acetyl-1,2,3,4-tetrahydroisoquinoline-3-carboxylic acid, please see:
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The indolecarboxylic acids are either from a commercial source or made from hydrolysis of corresponding esters or oxidation of corresponding aldehydes. The starting material for 13 is from Boc protection of 6-aminopenicillanic acid. For preparation of the starting material of 14, 2-acetyl-1,2,3,4-tetrahydroisoquinoline-3-carboxylic acid, please see:. Grunewald G.L., Romero F.A., and Criscione K.R. J. Med. Chem. 48 (2005) 134
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53349147286
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note
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2. Each condition was set up in triplicate and antiviral assays were performed independently on at least two separate occasions. Cell viability was measured 48 h post infection using the CellTiter-Glo Luminescent Cell Viability Assay (Promega), according to manufacturer's recommendations. Cell viability for the CellTiter-Glo Luminescent Cell Viability Assay was measured as luminescence and output expressed as relative luciferase units (RLU).
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23
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53349164642
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note
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Authentic SARS-CoV 3CLpro was diluted to 1 μM concentration using cold 20 mM HEPES pH 7.5. Five micrometers of compound 10 was added using a 1:400 dilution into the enzyme solution. The final volume was 200 μL and contained a final DMSO concentration of 0.25%. Control reactions were performed in the same buffer in the presence of 0.25% DMSO in the absence of inhibitor. Incubation was performed at room temperature for 20 min. Sinapinic acid was prepared as a saturating solution in 50% acetonitrile and 1% TFA. The sandwich method of preparation was used for MALDI. One microliters of the matrix was spotted onto a plate followed by 1 μL of the enzyme inhibitor complex followed by a top layering of 1 μL of matrix. A Voyager DE Pro Time of Flight (TOF) mass spectrometer was used for determining the molecular weights.
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24
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25
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