Convergence of retention for small solutes in reversed-phase liquid chromatography

Summary It is shown theoretically that when the concentration of organic solvent in the mobile phase increases, or solute size decreases, log k values of small solutes in reversed-phase liquid chromatography (RPLC) will tend to have a minimum value called the convergence point. A theoretical model for evaluating the convergent coordinates of small solutes is presented by using a stoichiometric displacement model for retention (SMDR). The physical meaning of the coordinates of each kind of convergence are also elucidated. The convergence points have either two-dimensional coordinates with a common ordinate (the logarithm of the phase ratio of the column, log ) or threedimensional corrdinates with two common axes: — log and the logarithm of the molar concentration of the pure displacing agent in mobile phase, log a_D. The other axis relates to the nature of the solutes, such as carbon number of a homolog, van der Waal's surface area, hydrophobic fragment constant etc. for the latter and those and/or concentration axis for the former. The model was tested with published data and found to give a good fit.     

Comparative study of some commercial chemically bonded phases in classical reversed-phase chromatography and in ion-pair reversed-phase liquid chromatography

Summary The behaviour of various octadecyl commercial bonded phases are compared in classical reversed-phase chromatography and in ion-pair reversed-phase chromatography. Great differences are exhibited by the packings studied according to the polarity of the solutes. Whereas hydrocarbonaceous bonded phases show very similar selectivity versus apolar or weakly polar solutes, great differences are observed when analyzing more polar solutes even when ion-pair reversed-phase chromatography is performed.Presented at the 14th International Symposium on Chromatography London, September, 1982     

反相离子对色谱法测定间苯二磺酸钠

运用反相高效离子对色谱法测定了间苯二磺酸钠.采用C18色谱柱(250 mm×4.6 mm i.d.,5 μm),流动相为V(乙腈):V(50 mmol/L NaH2PO4)＝7:93(含0.05%四丁基氢氧化铵),流速为1.0 mL/min,检测波长210 nm,柱温:30 ℃.间苯二磺酸钠在0.32×10-3～81.92×10-3 mg/mL范围内线性关系良好,r＝0.9999,加样回收率为98.8%～101.1%,RSD＝0.73%(n＝7).本法为工业生产间苯二磺酸钠提供了分析方法.     

Preparation and retention characteristics of different phenyl phases for reversed-phase liquid chromatography

Summary Three different phenyl phases were prepared. The amount of organic moiety bound on silica support was determined from thermogravimetric curves of the modified silica gel. The specific surface areas of gels were obtained from nitrogen sorption measurement at –196°C. The effect on separation and selectivity of the different bonded-phenyl functional groups was studied. The selectivity of the phenyl-bonded silica gels was examined and compared with octadecyl (ODS) silica in liquid chromatography. Corresponding to the high surface concentration of functional groups, the capacity factors of solutes, normalized to unit surface area of the adsorbent, k/S_BET were found to decrease in the sequence phenylmethyl>diphenyl>triphenyl. Polar solutes are retained in greater extent on the phenyl phases than on the ODS phase.     

Retention prediction of o-phthalaldehyde amino acid derivatives in reversed-phase liquid chromatography

Summary Retention prediction of o-phthalaldehyde amino acid derivatives in reversed-phase liquid chromatography has been investigated. The retention of all derivatives could be predicted within about 10% relative error under the appropriate separation conditions in both isocratic and gradient-elution modes.     

Retention prediction of small peptides in reversed-phase liquid chromatography

Summary Retention prediction of small peptides (up to four residues) in reversed-phase liquid chromatography has been investigated, considering the contributions of side chains in each position to the peptide retention. In isocratic elution the retention of peptides could be predicted within about 8% relative error.     

Retention prediction of phenylthiohydantoin amino acid derivatives in reversed-phase liquid chromatography

Summary The concept of retention prediction for the separation of phenylthiohydatoin-amino acid derivatives in isocratic reversed-phase liquid chromatography is described. A novel retention-solubility parameter, R, is defined, for the retention prediction strategy and the performance of this R value is evaluated by comparing measured and predicted retention data. Excellent agreement between these values were observed. It is concluded that the R value has a very high potential in describing the retention of phenylthiohydantopinamino acid derivatives withdifferent types of separation systems consisting of C-18, C-8 and phenethyl bonded stationary phases and various mobile phases.     

Interconversion of gradient and isocratic retention data in reversed-phase liquid chromatography: Effect of the uptake of eluent modifier on the retention of analytes

The experimental technique of mass spectrometric tracer pulse chromatography was used to study the effect of the sorption of eluent components by a C₁₈-bonded silica RPLC packing on the retention of a series of test analytes during isocratic and gradient elution experiments. The analytes of interest were a substituted phenol, a substituted nitroaniline, an anti-malaria drug, tetrahydrofuran, and methanol. The eluent used was a mixture of acetonitrile and water. The solutes and isotopically labeled eluent components were injected at fixed time intervals during each gradient run. The mass specific detector allowed the assignment of individual analyte peaks even when there was overlap in the chromatograms from successive injections. Thus, the retention time of each analyte could be determined as a function of gradient slope and initial eluent composition at the time of each injection. Experimental gradient retention time data were then compared with the calculated results from two theoretical models. The first model assumed the velocity of the mobile phase and eluent were equal. The second and most realistic model assumed the velocity of the eluent was less than the velocity of the mobile phase due to the uptake of eluent by the stationary phase. Gradient retention times predicted by the two models were reasonably accurate with the sorption model giving slightly more accurate values. Inverse calculations, i.e., calculation of isocratic retention factors from gradient elution data were also carried out with very similar results. That is, the model allowing for the uptake of eluent was slightly more accurate than the model assuming no eluent-stationary phase interaction.     

Optimization of reversed-phase ion-pair chromatography by over-pressurized thin-layer chromatography III. Over-pressurized thin-layer chromatography using 10-camphor sulfonic acid as ion-pair reagent

Summary The use of over-pressurized thin-layer chromatography in ion pair system using acidic type pairing reagent has been studied. The most important aspect when reversed phase ion-pair TLC system is used to apply a suitable procedure for pre-treatment of the sorbents. Because of the acidic type of ion pair reagents cannot be bounded to the surface of the layer by immersion or pre-development with the reagent solution, double coating technique was used for the pre-treatment, the plate was firstly immersed in an ethanolic solution of cetrimide, then the immersion has been repeated by an ethanolic solution of acidic ion pair reagent. The necessary coating of the sorbents can be achieved by this technique. To find the optimal conditions for reversed phase ion pair TLC separation of organic amines, 10-camphor sulfonic acid as reagent, different aliphatic, aromatic amines and diamines and heterocyclic nitrogen compounds, respectively as model compounds were selected. The dependence of the selectivity and efficiency of the separation on the sorbents, on the concentrations of reagents (cetrimide and camphor sulfonic acid) applied for both immersion and in the eluent were investigated in detail. Presented at the 15th International Symposium on Chromatography, Nürnberg, October 1984     

Peak-splitting in reversed-phase,ion-pair high-performance liquid chromatography of sympathomimetic drugs and its probable mechanism

Summary In the determination of ephedrine using reversed-phase, ion-pair liquid chromatography, a chromatographically pure sample was observed to give three peaks under certain mobile phase conditions. The mobile phases which produced maximum peak splitting were determined for ephedrine and a number of other sympathomimetic drugs.A proposal that peak splitting was the result of the composite interplay of two discrete chromatographic mechanisms, was investigated. The results of analysis by GC/MS confirmed that each peak was due to ephedrine, however, only one of the three split peaks was found to contain ion pairs. It is postulated that peak splitting is a physical phenomenon on reversed-phase columns and the separation of these drugs by ion-pair HPLC is based on a mixed rather than a single mechanism.This study has also shown that errors can arise in ion-pair HPLC when multiple peaks are assumed to indicate heterogeneity. Interconvertible species of the same solute can give rise to these peaks.     

Analysis of nucleotides,nucleosides, nucleobases in cells by ion-pair reversed-phase HPLC

Summary HPLC methods for the separation of nucleotides, nucleosides and nucleobases by ion-pair reversed-phase are reviewed. The advantages of these are discussed versus anion-exchange and reversed phase separations. Extraction procedures for nucleotide determinations from cells and tissues are pointed out in detail. Extracts from red blood cells, Ehrlich ascites tumour cells, hepatocytes, intestine are used for determination of nucleotide concentrations by the methods described.     

On the chromatography mechanism of reversed-phase liquid chromatography

Summary An equation is derived which can describe how the retention of solutes is influenced by the composition of the mobile phase in reversed-phase liquid chromatography, the retention of solutes in alkyl bonded stationary phase regarded as the complexation between solute molecule and the active sites on the surface of the stationary phase. When the stationary phase is not fully saturated by the organic modifier, the activity of the active sites, the activity coefficient of the adsorbed solute as well as the activity coefficient of the solute in the mobile phase depend on the composition of the mobile phase. However, when the stationary phase is fully saturated, the composition of the mobile phase mainly influences the activity coefficient of the solute in the mobile phase. In addition, the selectivity of retention is discussed in terms of the derived equation.     

反相离子对色谱法测定脱氧精胍菌素药物中的主成分

建立了反相离子对色谱法分离测定脱氧精胍菌素药物中主成分的方法。分别考察了色谱柱类型、离子对试剂种类及浓度、缓冲盐浓度和流动相pH值等参数对实验结果的影响。确定了分离脱氧精胍菌素的最佳条件: C18色谱柱(250 mm×4.6 mm, 5 μm);以5 mmol/L磷酸氢二钾水溶液(含5 mmol/L戊烷磺酸钠,pH 3.6±0.3)-乙腈(90:10, v/v)为流动相,流速为1.0 mL/min;检测波长为210 nm;柱温为30 ℃;进样量为20 μL。该方法实现了对脱氧精胍菌素的良好分离,且线性关系良好,检出限可达0.5 mg/L。     

Computer interactive identification system for polycyclic aromatic hydrocarbons in reversed-phase microcolumn liquid chromatography

Summary A computer interactive identification system is proposed which is based on the relationship between retention and molecular properties such as the size and shape of polycyclic aromatic hydrocarbons (PAHs). This system offers an automatic analytical process for liquid chromatography, providing a reliable identification of the separated components. The identification can be further enhanced by the use of multiple detectors such as a multichannel UV detector. The system can be used for optimization procedures, resulting in a highly automatic complex analytical system.     

Separation of inorganic anions by reversed-phase ion-pair chromatography with direct UV detection

In this paper, a separation method of the inorganic anions including I^–, NO ₂ ^– , NO ₃ ^– , IO ₃ ^– and SCN^– on the reversed-phase ion-pair chromatography with direct UV detection has been developed, and the limits of detection of these inorganic anions were determined. The effects of the organic modifier volume fraction and concentration of the ion-pair reagent on the retention of inorganic anions were discussed.     

Determination of oxybutinin chloride in pharmaceuticals by reversed-phase ion-pair liquid chromatography with two counter-ions in the eluent

A reliable stability-indicating method using reversed-phase ion-pair high-performance liquid chromatography for the determination of the anticholinergic drug oxybutinin chloride in pharmaceuticals is reported. Sample extraction is easy and fairly rapid and recovery and precision of the method are excellent. Due to the simultaneous use of an organic amine and of an alkanesulphonate in the mobile phase, good selectivity towards related (quaternary ammonium) anticholinergic drugs was obtained.     

Simple models for the effect of aliphatic alcohol additives on the retention in reversed-phase liquid chromatography

Four retention models for the effect of aliphatic alcohol additives on the retention of analytes in reversed-phase liquid chromatography have been developed following either a semi-thermodynamic treatment or an empirical approach. Their performance was tested using the experimental retention times of six non-polar analytes (alkylbenzenes) and ten o-phthalaldehyde derivatives of amino acids under different isocratic chromatographic runs when a small amount of ethanol, 1-propanol, 1-butanol, 1-pentanol, 1-hexanol or 1-heptanol was added to methanol/water mixtures containing a constant amount of methanol. It was shown that for the structurally simple alkylbenzenes all the models can be adopted for retention prediction with good results. In contrast, just one out of four models, that with the fewest approximations, predicts satisfactorily the retention properties of amino acids derivatives. However, the most interesting feature is that this model can predict the effect of an alcohol-additive on the retention properties of solutes, even if this additive was not used in chromatographic runs done for the fitting procedure, provided that it belongs to the same homologous series of alkanols. This feature is also observed in all models described the retention of alkylbenzenes.