Effect of pH on UV Absorbance

UV spectra of aromatic hydrocarbons are characterized by three sets of bands (El, E2, and B bands) that originate from ti ti transitions. Generally the E2 and B bands are of most interest to chromatographers, since the solvent cutoff for most mobile phases is 200 nm. 

TABLE 8-9. Molar Absorptivity Values for Neutral, Acid, aud Basic Species  

Effect of pH on UV absorbance for aniline (obtained from diode array). v/v% acetonitrile and pH of 15 mM K2HPO4 7H2O adjusted to pH 1-9 with (B) pH2.0 and acetonitrile concentration changed from 10 to 50 v/v%. 

In order to avoid any secondary equilibrium effects on the retention of iono-genic analytes, it is preferable to use a mobile-phase pH either two units greater or less than the analyte pA. Therefore knowledge of the analyte pA is very important. A basic understanding of how functional group substitution on a molecule affects the pA of the ionizable group on the substrate is given. An exhaustive description of all the nuances of analyte substitution on analyte pAa is not included in this section. However, further details can be found in the references 29-31. 

Effect of Analyte Substitution on Analyte pKa. The acidity of substituted phenols or carboxylic acids depend upon the substituent attached to the cor- 

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Figure 8-57. Effect of pH on UV absorbance for aniline. Conditions Column 15-cm X 0.46-cm Luna C18(2). Eluent 90% aqueous 10% MeCN. Aqueous 15mM K2HP04-7H20 adjusted to "pH 1.5-9 with H3PO4, Flow rate, ImL/min temperature,...

Fig. 14. Effects of temperature on the absorbance of hemopexin and the N-domain of hemopexin. The unfolding of hemopexin and N-domain in 25 mM sodium phosphate, pH 7.4, was examined using absorbance spectroscopy (N. Shipulina et al., unpublished). The second derivative UV absorbance spectra of the protein moieties were used to follow protein unfolding and the Soret and visible region spectra to monitor the integrity of the heme complexes, as done with cytochrome 6502 (166). The ferri-heme complex is more stable than the apo-protein moiety, but the is slightly lower than that assessed by DSC, indicating that changes in conformation occur before thermodynamic unfolding. Reduction causes a large decrease in heme-complex stabihty, which is proposed to be a major factor in heme release from hemopexin by its cell membrane receptor, and addition of 150 mM sodium chloride enhanced the stabihty of ah forms of hemopexin.

Fig. 7 Effect of pH on the surface plasma resonance absorbance on UV-vis spectra of Au-CAH/ CMC aqueous solution. Inset photos show the color of the Au-CAH/CMC dispersions at different pH (Reprinted, with permission, from [228], Copyright (2010) American Chemical Society)...

The nonenzymatic browning reactions of fructose and fructose-lysine aqueous model systems were investigated at 100°C between pH 4.0 and pH 12.0 by measuring the loss of reactants and monitoring the pattern of UV-absorbance and brown color development [2]. At all the pH values tested, the loss of fructose was lower in the presence than in the absence of lysine. The promoting effect of pH was clear on the browning development and was in agreement with the earlier studies [4,197],... 

The effect of solution chemistry on the speciation of the organic contaminant 1-naphtol (1-hydroxynaphthalene) and its complexatiom with humic acid is reported by Karthikeyan and Chorover (2000). The complexation of 1-naphtol with humic acid (HA) was studied during seven days of contact, as a function of pH (4 to 11), ionic strength (0.001 and 0.1 M LiCl), and dissolved concentration (DO of 0 and 8 mg L ) using fluorescence, UV absorbance, and equilibrium dialysis techniques. In a LiCl solution, even in the absence of HA, oxidative transformation of 1-naphtol mediated by was observed. In addition, the presence of humic acid in solution, in the absence of DO, was found to promote 1-naphtol oxidation. These reactions are affected by the solution chemistry (pH, ionic strength, and cation composition). 

Shen et al. (1995) investigated the effect of light absorbance on the decomposition of aqueous chlorophenols (CPs) by UV/H202. The photoreaction system was batch annular photoreactors with 254-nm, low-pressure UV lamps at 25°C. The light absorbance and photolytic properties of chlorophenols and H202 were found to be highly dependent on the solution pH and can be adequately described with the linear summation of the light absorbance of undissociated and dissociated species of chlorophenols ... 

Figure S.3 Effect of capillary length on the resolution of tryptic fragments of BSA. Conditions capillary, 375 /xm O.D. X 50 /xm I.D. buffer, 100 mM sodium dihydrogen phosphate, pH 2.5 temperature, 25°C detection, UV absorbance at 200 nm injection, pressure for 10 sec sample, 10/1 dilution of BSA digest. (Reprinted from Ref. 5 with permission.)...

A method for determination of sodium isoascorbate (see 2) in boiler feed water, where it is used for deoxygenation, consists of following the reaction kinetics of Rhodamine B (13) in the presence of KBrOs, measuring at 555 nm. A linear correlation exists between the catalytic effect of the analyte on the reaction rate and its concentration Fe(III), Ca(II) and Mg(II) in the 5-200 ppm range interfere with the analysis . The effects of solvents, pH, surfactants, metal ions and other food additives on the absorbance were studied for the micelle-enhanced UV spectrophotometric determination of the food preservative sodium D-isoascorbate. The optimal conditions were using water at pH 7-8 as solvent and polyvinyl alcohol as surfactant, which causes an up to 3-fold increase of the UV absorbance. ... 

Dissolved humic and fulvic acids can be isolated from the total DOC pool by adsorption onto resins as discussed by Aiken in Chapter 14 of this book. Muenster (1982) found that 77-86% of the DOC (measured by UV absorbance and organic carbon) adsorbed onto XAD resin at pH values of 2.0-3.0. Adsorption onto PVP resin was slightly less effective (70-76% at pH values of 2.0-3.0). These differences in adsorption efficiency may be explained by organic material other than humic substances being retained on these resins. 

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