Tryptophan light absorption

Comparison of the light absorption spectra of the aromatic amino acids tryptophan and tyrosine at pH 6.0. The amino acids are present in equimolar amounts (10 3 m) under identical conditions. The measured absorbance of tryptophan is as much as four times that of tyrosine. Note that the maximum light absorption for both tryptophan and tyrosine occurs near a wavelength of 280 nm. Light absorption by the third aromatic amino acid, phenylalanine (not shown), generally contributes little to the spectroscopic properties of proteins. 

Spectra of proteins and nucleic acids. Most proteins have a strong light absorption band at 280 nm (35,700 cm ) which arises from the aromatic amino acids tryptophan, tyrosine, and phenylalanine (Fig. 3-14). The spectrum of phenylalanine resembles that of toluene (Fig. 23-7)whose 0-0 band comes at 37.32 x 10 cm. The vibrational structure of phenylalanine can be seen readily in the spectra of many proteins (e.g., see Fig. 23-llA). The spectrum of tyrosine is also similar (Fig. 3-13), but the 0-0 peak is shifted to a lower energy of 35,500 cm (in water). Progressions with spacings of 1200 and 800 cm are prominent. The low-energy band of tryptophan consists of two overlapping transitions and The Lb transition has well-resolved vibrational subbands, whereas those of the La transition are more diffuse. Tryptophan derivatives in hydrocarbon solvents show 0-0 bands for both of these transitions at approximately... 

For gel filtration experiments buffer A is used, which contains additionally 200 pM of the respective nucleotide. Prior to injection of 200 p of 20 pM hGBPl, the column is equilibrated with two column volumes of this buffer. The eluting protein is detected by UV absorption or, more sensitively, tryptophan fluorescence. A wavelength of 295 nm should be selected for detection or excitation, respectively, because the nucleotide in the buffer leads to strong light absorption at lower wavelengths. 

The appearance and disappearance of the triplet state can be measured by light emission or by absorption change. The absorption change arises because the ground and triplet states have different absorption spectra. The absorption spectrum of tryptophan in the triplet state is red shifted in com-... 

A number of investigators have studied the effect of ozone on the ultraviolet absorption spectra of proteins and amino acids. A decrease in the absorption of 280-nm light in a number of proteins was originally reported ly Giese et aV to be a consequence of ozone exposure they suggested that this was due to an interaction of ozone with the ring structures of tyrosine and tryptophan. Exposure of a solution of tryptophan to ozone resulted in a decrease in 280-nm absorption, whereas the extinction coefficient of tyrosine increased. Similar results with tyrosine were reported by Scheel et who also noted alterations in the ultraviolet spectra of egg albumen, perhaps representing denaturation by ozone. 

Side chains of the three aromatic amino acids phenylalanine, tyrosine, and tryptophan absorb ultraviolet light in the 240- to 300-nm region, while histidine and cystine absorb to a lesser extent. Figure 3-13 shows the absorption spectrum of a "reference compound" for tyrosine. There are three major absorption bands, the first one at 275 nm being a contributor to the well-... 

Determination of protein concentration by measuri ng absorbance at 280 nm (A2g0) is based on the absorbance of UV light by the aromatic amino acids tryptophan and tyrosine, and by cystine, disulfide bonded cysteine residues, in protein solutions. The measured absorbance of a protein sample solution is used to calculate the concentration either from its published absorptivity at 280 nm (a280) or by comparison with a calibration curve prepared from measurements with standard protein solutions. This assay can be used to quantitate solutions with protein concentrations of 20 to 3000 pg/ml. 

Absorption spectrum is the plot of light intensity as a function of wavelength. Figure 1.2 shows the absorption spectra of tryptophan, tyrosine, and phenylalanine in water. A strong band at 210-220 nm and a weaker band at 260-280 nm can be seen. 

Among the properties of amino adds that are most pertinent to the biomedical scientist are their optical rotations, already discussed, which are listed for each amino acid in Table 4.1. Note the dramatic differences between optical rotations in the zwitterionic (water) and fully protonated (HC1) forms. Further, all amino acids absorb ultraviolet light in the range 190-220 nm. The C=0 bond in carboxyl residues is largely responsible. Moreover, aromatic amino acids, especially tryptophan, absorb in the 260-285 nm range. Protein concentrations in solutions are often determined via absorption at 210 or 280 nm. 

The part of a molecule that absorbs the light and is, therefore, responsible for its colour (whether in the visible or UV region) is called the chromophore, and the wavelength dependence of the absorption defines its absorption spectrum. Figure 7-3 illustrates the absorption spectrum of the three aromatic amino acids tryptophan, tyrosine and phenylalanine. 

This chapter is devoted to describe the impact of metallic nanosphere to the multi-photon excitation fluorescence of Tryptophan, and little further consideration to multi-photon absorption process will be given, as the reader can find several studies in [11-14]. In section II, the nonlinear light-matter interaction in composite materials is discussed through the mechanism of nonlinear susceptibilities. In section III, experimental results of fluorescence induced by multi-photon absorption in Tryptophan are reported and analyzed. Section IV described the main results of this chapter, which is the effect of metallic nanoparticles on the fluorescent emission of the Tryptophan excited by a multi-photon process. Influence of nanoparticle concentration on the Tryptophan-silver colloids is observed and discussed based coi a nonlinear generalization of the Maxwell Garnett model, introduced in section II. The main conclusion of the chapter is given in secticHi IV. 

Due to its relevance to the next section, we observed and analyzed the fluorescent emission of Tryptophan in water solution excited by one, two, and three-photon absorption. For that, three different light sources were used a UV (180-375 nm) lamp, the second harmonic of a Q-switched Nd YAG laser (with 8 ns pulse duration at 532 nm) and a Ti-Sapphire laser delivering pulses at 76 MHz, with 150 fs pulse duration and 500 mW average power at 800 nm. 

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