Solutes, chromatographic behavior

Method of Moments The first step in the analysis of chromatographic systems is often a characterization of the column response to sm l pulse injections of a solute under trace conditions in the Henry s law limit. For such conditions, the statistical moments of the response peak are used to characterize the chromatographic behavior. Such an approach is generally preferable to other descriptions of peak properties which are specific to Gaussian behavior, since the statisfical moments are directly correlated to eqmlibrium and dispersion parameters. Useful references are Schneider and Smith [AJChP J., 14, 762 (1968)], Suzuki and Smith [Chem. Eng. ScL, 26, 221 (1971)], and Carbonell et al. [Chem. Eng. Sci., 9, 115 (1975) 16, 221 (1978)]. 

Because trls has only meager buffering action at pH 6.1, fluctuations In the pH of Ion exchanger and solutions may occur and may result In abnormal chromatographic behavior. Buffered conditions may be obtained by substituting bls-trls which has a pK of about 6.5, and virtually Identical behavior results If Developer A Is constituted with 0.03 M bls-trls-HCl 0.03 M NaCl 0.01% KCN at pH 6.2. 

The mobile phase should not produce chemical transformations of the separated components because it can modify the chromatographic behavior of the system. Solvents having weak reversible bonds with the solute are recommended. 

The different organic modifiers used to derive the most suitable mobile phases lead to different parameters namely isocratic logfe and extrapolated logkw. The extrapolation method has no reality in terms of chromatographic behavior of solutes. However, mainly by correlation with log Pod (Eqs. 2 and 3) several studies have demonstrated the interest of these extrapolated retention factors as predictors of the lipophilicity of solutes. 

To identify isomeric thienothiophenes, the chromatographic behavior of mono- and dialkyl-substituted thienothiophenes 1 and 2 was studied.Thienothiophene 1 and its alkylated derivatives were shown to be characterized by greater retention volumes than the corresponding thienothiophenes 2. The linearity of the retention volume vs. boiling point relationship allowed the thienothiophene isomers to be identified. Studies on solution thermodynamics of thienothiophenes in the stationary phase showed that isomeric thienothiophenes 1 and 2 do not differ appreciably in their heats of solution. For example, the calculated heats of solution of 5-ethyl-3-methylthieno[2,3-h]thiophene (26) and 5-ethyl-3-methylthieno[3,2-b]thiophene (27) in polyethyleneglycol adipate are both about 16 kcWmole. ... 

Adsorption. Hydrophobic interactions, which may occur using aqueous mobile phases, usually can be eliminated by the addition of an organic modifier to the aqueous mobile phase (30,33) or by a reduction of ionic strength (3A 25.)- Recently, Haglund and Marsden (36-AO) have undertaken a systematic study on the chromatographic behavior of low molecular weight solutes on Sephadex packings and explained these results in terms of hydrophobic interactions. 

Relative humidity and temperature are two variables which have influence on the chromatographic behavior of the solutes [2] but which can not always be set at desired levels. The relative humidity is expected to have a large influence, while temperature has a small influence. In reference [2] it is stated that a temperature change of 5 degrees seldom exceeds reproducibility limits of the standard working techniques. It is most feasible to discuss the effect of variation in relative humidity and temperature in terms of activity. Therefore in the following paragraphs first the concept of activity will be introduced. Then the concept will be applied in a short examination of the effect of relative humidity and temperature on the retention. 

Although HETP is a useful concept, it is an empirical factor. Since plate theory does not explain the mechanism that determines these factors, we must use a more sophisticated approach, the rate theory, to explain chromatographic behavior. Rate theory is based on such parameters as rate of mass transfer between stationary and mobile phases, diffusion rate of solute along the column, carrier gas flowrate, and the hydrodynamics of the mobile phase. 

The term interfering solutes has been coined to characterize this situation. The seemingly minor complication has serious consequences for the theoretical treatment and may be said to add a new dimension to chromatographic behavior. An examination of the effects caused by solute interference is the subject of the present communication. 

These amino-acid derivatives can be separated from the ordinary amino acids resulting from hydrolysis of the peptide because the low basicity of the 2,4-dinitrophenyl-substituted nitrogen (Section 23-7C) greatly reduces the solubility of the compound in acid solution and alters its chromatographic behavior. The main disadvantage to the method is that the entire peptide must be destroyed in order to identify the one A-terminal acid. 

The sample diluent affects the solute dispersion. If we consider the effects of three different diluents (hexane, chloroform, and acetone) on the chromatographic behavior of a TG mixture on RP columns using, for example, acetonitrile and ethanol as the mobile phase, we can see that the TGs dissolved in hexane provided only a minute chromatographic trace, whereas dissolution in chloroform yielded excellent detection and resolution. These results can best be explained by invoking the solvophobic theory of Horvath and Melander (85). 

With binary and ternary supercritical mixtures as chromatographic mobile phases, solute retention mechanisms are unclear. Polar modifiers produce a nonlinear relationship between the log of solute partition ratios (k ) and the percentage of modifier in the mobile phase. The only form of liquid chromatography (LC) that produces non-linear retention is liquid-solid adsorption chromatography (LSC) where the retention of solutes follows the adsorption isotherm of the polar modifier (6). Recent measurements confirm that extensive adsorption of both carbon dioxide (7,8) and methanol (8,9) occurs from supercritical methanol/carbon dioxide mixtures. Although extensive adsorption of mobile phase components clearly occurs, a classic adsorption mechanism does not appear to describe chromatographic behavior of polar solutes in packed column SFC. 

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