Phenol resonance energy

Figure 10.12. Absorption spectra of Phenol Red and emission spectrum of Eosin as potential pH probe based on resonance energy transfer.

INTRINSIC AND EXTRINSIC FLUORESCENCE. Intrinsic fluorescence refers to the fluorescence of the macromolecule itself, and in the case of proteins this typically involves emission from tyrosinyl and tryptopha-nyl residues, with the latter dominating if excitation is carried out at 280 nm. The distance for tyrosine-to-tryp-tophan resonance energy transfer is approximately 14 A, suggesting that this mode of tyrosine fluorescence quenching should occur efficiently in most proteins. Moreover, tyrosine fluorescence is quenched whenever nearby bases (such as carboxylate anions) accept the phenolic proton of tyrosine during the excited state lifetime. To examine tryptophan fluorescence only, one typically excites at 295 nm, where tyrosine weakly absorbs. [Note While the phenolate ion of tyrosine absorbs around 293 nm, its high pXa of 10-11 in proteins typically renders its concentration too low to be of practical concern.] The tryptophan emission is maximal at 340-350 nm, depending on the local environment around this intrinsic fluorophore. 

The keto tautomer of phenol does not retain any of the resonance energy associated with the aromatic ring, while the keto tautomer of 2-hydroxypyridine does (compare with amide resonance). 

The effect of rf coordination on the arenes was studied in the context of the phe-nol-ketodiene equilibrium . It was shown that this equilibrium for the free ligands favors heavily the phenol tautomer vide supra) whereas for the complexes [Os(NH3)5-2,3- -arene)] + (arene = phenol 2-, 3-, 4-methylphenol 3,4-dimethylphenol) the corresponding equilibrium constants approach unity (20 °C). The conversion of phenol 67 into the 2,4-cyclohexadien-l-one 68 was kinetically favored over the formation of the 2,5-isomer 69, although the latter is the thermodynamically favored product (equation 26). It was assumed that osmium rehybridizes the C(5) and C(6) atoms to form a metallocy-clopropane. This removes much of the resonance energy and therefore destabilizes the enohc form of the free hgand. The free energies of ketonization (25 °C) for the /j -phenol complex in comparison with free phenol are shown in equations 27 and 28 . 

Resonance energies, in keto/enol forms of phenol 39... 

Electron delocalization can affect the nature of the product formed in a reaction and the of a compound. A carboxylic acid and a phenol are more acidic than an alcohol such as ethanol, and a protonated aniline is more acidic than a protonated amine because electron withdrawal stabilizes their conjugate bases and the loss of a proton is accompanied by an increase in resonance energy. 

Here, we are dealing specifically with keto-enol tautomerism (Figure 6.17). Generally, in aliphatic compounds, the keto form is in great predominance for example, in acetone less than 10 % is in the enol form. At the other extreme is the phenolic stmcture, where there is no evidence for the existence of the keto form. This would not be unexpected because a loss of the resonance energy of stabilization would be entailed. 

AG° values for the aromatic ring of phenol on the basis of tautomeric equilibrium constants. When AG° values can be converted into Aff values, the resonance energy differences between two structures can be determined. The value found for the benzene aromatic resonance energy (32 kcal/mol) is in agreement with other experimental and calculated data. 

The indole group of tryptophan (Trp) has a higher molar extinction coefficient than the phenolic and phenyl side chains of tyrosine (Tyr) and phenylalanine (Phe), respectively (see Table 1, Chapter 12). Thus although its quantum yield is similar to that of lyr, its fluorescence emission is much mote intense. Furthermore, its excitation spectrum overlaps the emission spectrum of Tyr and therefore fluorescence resonance energy transfer (FRET, see Chapters 2 and 3) from lyr to Trp occurs readily when both residues are in close proximity (i.e. located in the same protein molecule) and favourably orientated. The intrinsic fluorescence of a protein is therefore dominated by the contribution from Trp... 

The resonance energy for these five structures stabilizes the phenolate ion more than the amount by which the undissociated phenol molecule is stabilized by resonance between the two Kekule structures (with only small contributions by the other three, which involve a separation of charges). The extra stabilization of the anion increases the acid constant the observed factor 10" corresponds to the reasonable value 33 kJ moIe for the extra resonance energy of the phenolate ion. 

The nuclear magnetic resonance spectra of SchifF bases formed from primary amines and ortAo-hydroxy aldehydes and ketones show that the Schiff bases derived from l-hydroxy-2-acetonaphthone and from 2-hydroxy-1-naphthaldehyde exist as keto amines (7a) although their formation involves loss of most of the resonance energy of one of the aromatic rings When R is a phenyl group, the phenol-imine tautomer (7b) predominates Schiff bases derived from ortho-hydroxy aldehydes and ketones have the phenol-imine structure (8) . Evidently, in such compounds the keto-amine tautomer... 

Lysine and Tyr side chains can quench tryptophan fluorescence by transferring a proton to the excited indole ring [59, 75]. The phenolate side chain of ionized tyrosine also can quench by resonance energy transfer and possibly by transferring an electron to the indole [51]. 

The result of this resonance stabilization of the anion by delocalization of the negative charge makes the overall energy change required for ionization to be smaller for phenol than it is for ethanol. This can be shown in terms of energies as follows ... 

Antioxidants have to form stable low-energy free radicals that will not further propagate the oxidation of fats and oils. From this point of view, the most convenient compounds are the phenolic compounds, which structure allows them to form low-energy radicals through stable resonance hybrids, as follows ... 

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