甲胺磷金属配合物中主要成分含量检测方案

Application Note Advanced application of CD measurement for pharmaceutical researchAnalyzing the interaction of human serum albumin and 3，5-diiodosalicylic acid2/3 Advanced application of CDmeasurementfor pharmaceutical research Analyzing the interaction of human serum albumin and 3，5-diiodosalicylic acid Introduction Human serum albumin (HSA) is the mostabundant protein in blood plasma. HSA bindswith pharmaceutical compounds and other in vivosubstances， and plays an important role in thetransport of these substances to target organs. Severalstudies have reported on the binding affinity of HSAand its interaction with various compounds 1)-3). Herein， the measurement of CD induced by theinteraction of HSA titrated with 3，5-diiodosalicylic acidand subsequent analysis of the dissociation constant ispresented. Keywords Proteins， Induced CD， Pharmaceuticals JASCO J-1500 CD Spectrometer Application Note Measurement Conditions Measurement System Sample： HSA， 0.0228mM， 100 mMacetate buffer (pH 6.3)， 2 J-1500 CD Spectrometer mL PTC-517 Peltier thermostatted cell holder Titration： 3，5-diiodosalicylic acid， (0.25 mM)， 50 mL， 20 ATS-522 Automatic titrator times Scanning range： 360 to 260 nm Results Chirality resulting from an achiral substance interacting with a chiral substance is called induced CD. Though，3，5-diiodosalicylic acid is achiral， it exhibits chirality through interaction with HSA. The resulting CD spectrashow a positive peak around 320 nm. (Fig. 1) Fig. 1 3D-CD spectra of HSA titrated with 3，5-diiodosalicylic acid HSA does not show a CD signal around 320 nm， but the induced CD from the interaction of HSA and3，5-diiodosalicylic acid increases with additional titration of 3，5-diiodosalicylic acid. (Fig. 2) ( A pplication L i brary： http：//www.jascoinc.com/applications ) Fig. 2 2D-CD spectra of HSA titrated with 3，5-diiodosalicylic acid The arrow represents increasing volume of 3，5-diiodosalicylic acid A plot of the increase of the induced CD signal (at 320 nm) versus the increase in 3，5-diiodosalicylic acid (Fig.3) and the Hill plot (Fig. 4) are shown below. 2 v=3.1386x+5.1385 1.5- 1 0.5 0 -1 80 -0.50 1 1.5- log[L] (mM) Fig. 3 Increase of induced CD (320 nm) Fig. 4 Hill plot The estimated dissociation constant (Kd)， which is the concentration at which 50% of the 3，5-diiodosalicylicacid is bound with HSA is 0.023 mM. The Hill coefficient is approximately 3.1， indicating a positive cooperativereaction. ( ) (1) R. Brodersen，J. Biol. Chem. (1977) 252，14，5067-5072 (2) UIrich KRAGH-HANSEN， Biochem. J.(1983) 209，135-142 (3) S. S. Sinha， R. K. Mitra and S. K. Pal， J. Phys. Chem. B. (2008)112， 4884-4891 ( A pplication L i brary： http：//www.jascoinc.com/applications ) JASCO INC.Mary's Court， Easton， MD USATel： ( Fax：( pplication Library： http：//www.jascoinc.com/applications IntroductionCD spectra measurement can be used forspectra measurement of organic compounds andmetalcomplexes.In this note, we show several CD spectra data