Copper Flavonol Complexes as Models for QD Presenting Author: P. Sisco Co-Authors: M. Perkovic*, D. ivens, W. Lynch AASU, Chemistry and Physics Dept. Savannah GA *Western Michigan University, Chemistry Dept., Kalamazoo, MI
Biomimic Chemistry Small molecule synthesis used to prepare relevant molecules Small Inorganic Molecules Serve as Structural and Functional Models Give insight into active site structure of biomolecules Provide chanistic Details Perform other functions Catalytic transformations Synthesis
Introduction Quercetin is a flavanoid compound in plants Quercetin 2,3-dioxygenase (QD) is a widely occurring copper metalloenzyme in fungi QD catalyzes ring opening cleavage of quercetin with molecular oxygen Recent literature presented a crystal structure and active site conformation for QD purified from Aspergillus japonicus Results of our studies of a small molecule inorganic biomimic of QD are presented Potential for future interest (area of bioremediation)
Proposed Mode of xidation Can. J. Chem., 1977, 16, 493.
Experimental Approach Tris-1-ethyl-4-methylimidazolylphosphine (T14MIP) Copper (II) Complexes P Cu 2+ + H 2 CH 1+ P Cu 2 CH
Copper - Flavonol Complex Preparation Scheme P Cu + + 2 H X X = H,,, Cl P Cu X 1+
Substituted Flavonols Experimental Approach H X 1. ah 2. ah, H 3. H 2 2 X H X = Cl,,
Substituted Flavonols [Cu(T14MIP)(flav-X)] + X UV-Vis(nm) Fluorescence nm emission, nm 440 (24,400) 538 425 (21,200) 440 (32,000) 536 422 (27,200) Cl 434 (25,200) 528 417 (20,900)
Xray Structure of Cu(T14MIP)(flav) + (3) C(10) C(11) C(6) C(5) C(4) C(12) C(28) C(7) C(3) C(2) C(1) (1) Cu(1) (2) C(13) C(8) C(9) C(14) C(16) C(15) (6) C(29) (1) C(17) C(30) C(22) C(23) (5) C(31) C(32) 33) C(25) C(24) P(1) (4) C(26) (3) C(18) C(19) (2) C(20) C(21) 7) Synthesis: CuX, T14MIP, 3-hydroxyflavone
Xray Table Cu(T14MIP)(flav) + Cu Bond Distances (Å) Protein/Substrate Predicted (Å) (1) - Cu(1) 2.013(7) Cu-ε2 (His66) 2.2 (3) - Cu(1) 2.269(8) Cu-ε2 (His68) 2.1 (6) - Cu(1) 1.998(7) Cu- ε2 (His112) 2.1 (1) - Cu(1) 2.022(7) (2) - Cu(1) 1.939(7) Cu Bond Angles ( ) (1) - Cu(1) - (3) 91.6(3) (1) - Cu(1) - (6) 89.3(3) (1) - Cu(1) - (1) 171.9(3) (1) - Cu(1) - (2) 95.8(3) (3) - Cu(1) - (6) 91.0(3) (3) - Cu(1) - (1) 96.5(3) (3) - Cu(1) - (2) 102.8(3) (6) - Cu(1) - (1) 90.2(3) (6) - Cu(1) - (2) 165.1(3) (1) - Cu(1) - (2) 82.7(3) τ =0.11 (implies near tetragonal geometry)
Functional Studies Structural model complexes have been prepared Spectroscopic evidence indicates that flavanol chemsitry can be easily monitored ne such spectrosopic technique is fluorescence as flavanols are known to be fluorophores Functional Studies Include Thermal and Light Initiated Degradation Reactions followed and Characterized by Fluorescence UV-VIS GC/MS
Cu(T14MIP)(flav)PF 6 Solid State Fluorescence Spectra 1100 900 Fluorescence (arbitrary units) 700 500 300 100 340 440 540 640 740-100 Wavelength (nm) Free 3-hydroxyflavone (solid state) excitation at 480 nm emission at 530 nm Complex (solid state) excitation at 431 nm emission, 695 nm and 718 nm Excitation Spectra collected with emission at 644 nm Emission spectra collected using excitation at 431 nm
Dissociation of 3-Hydroxyflavonate P Cu 1+ P Cu 2+ + - λ-max= 431 nm λ-max= 343 nm Experimental Parameters: 1. Acetonitrile or DMF 70 C 2. CH 2 Cl 2, RT, 431 nm
Dissociation of Cu(T14MIP)(flav)Cl at Elevated Temperature 1 0.8 Abs 0.6 0.4 Series1 Series2 Series3 Series4 0.2 0 300 400 500 nm 2.0 X 10-4 M Cu(T14MIP)(flav)chloride 70 C, DMF Series 1-4: 0 min., 10 min., 30 min., 1 h
Cu(T14MIP)(flav)PF 6 Light Promoted Dissociation 8 y = 0.0057x 2-0.3905x + 6.9486 R 2 = 0.989 Ratio (431nm/343nm) 6 4 2 0 0 10 20 30 40 Time(min) Dissociation using Visible Light at 431 nm Monitor decrease in Absorbance at 431 nm with corresponding increase in Absorbance at 343 nm 1.4 1.2 1 0.8 0.6 0.4 Time 1 Time 2 Time 3 Time 5 Time 6 0.2 0 280 330 380 430 480 Wavlength(nm) Isobestic Point at 373 nm
Degradation Products of 3-hydroxyflavonate - 2 CH H 2 A B C- C- C -Products esterified with diazomethane -Confirmed by GC-MS vs internal std. -Reported as mole equivalents
Degradation Products of 3-hydroxyflavonate marked with 18-2 CH H 2 A B C- C- C currentp 310 nm excitation* A B C 1.01 0.28 0.26 Thermal decomposition (120 C)** A B C 0.51 0.25 0.34
Degradation Products of Substituted 3-hydroxyflavonate X X - 2 CH X H A C- B C- X A (mole eq.) B(mole eq.) Cl - 0.21-0.25-0.33
Cu(T14MIP)(flav)PF 6 UV-Light Degradation y = 0.0031x 2-0.4398x + 16.568 R 2 = 0.9916 20 Ratio (431nm/343nm) 15 10 5 0 0 20 40 60 80 Time (min) Degradation Using UV-light at 300nm. Degradation monitored by decrease in Absorbance at 431 nm 1.2 0.8 0.4 Time 1 Time 2 Time 3 Time 4 Time 5 0 280 330 380 430 480 Wavelength(nm)
Degradation Products of 3-hydroxyflavonate P Cu 300 nm excitation* A B C 0.23 0.75 0.0 Thermal decomposition (120 C)** A B C 0.30 0.23 1.0 * Cl 4- salt in CH 2 Cl 2, ** in DMF Values are turnover number (substrate/copper) 1+
Degradation Products of 3-hydroxyflavonate P Cu 2+ + - Thermal decomposition (120 C) A B C 0.44 0.37 0.28 Thermal decomposition (120 C)* A B C 4.8 5.5 6.4 * in DMF 10 X 3-hydroxyflavonate Values are turnover number (substrate/copper)
Conclusions We have successfully prepared copper complexes which are biomimics of Quercetin Dioxygenase We have successfully prepared copper complexes with substituted flavonols to probe the electronic effects on the degradation process We have used spectroscopic techniques to determine the influence of electronic factors Degradation of flavonol can be monitored by GC/MS Degradation can be achieved thermally (120 C) or by 300 nm excitation. Dissociation can be achieved thermally (70 C) or by 431 nm excitation.
Acknowledgments AASU Department of Chemistry and Physics AASU Research and Scholarship Committee GC/MS SF-ILI DUE 9451465 MR SF-CCLI DUE 9952343 ACS - PRF 30550-GB3