Between 'Brush' and 'Bulk' Reverse Phases and Aqueous Solvents The interactions between aqueous solvents and brush reverse phases differ very significantly from those with a  bulk reverse phase at very low concentrations of solvent. This difference has been investigated by a number of workers (25-27) and the basic difference between the two types of phase are shown in the curves relating retention volume of methanol to the concentration of methanol in the mobile phase in figure 35. The phases shown are the RP-18 brush, reverse phase manufactured by E. M. Laboratories, which had a C18 (dimethyloctadecyl) chain and ODS-3 a bulk reverse phase which had a C18 (octadecyl) chain and was manufactured by Whatman Inc. The curves relating retention volume with solvent composition for the two phases show very different behavior patterns. The ODS-3 bulk reverse phase behaves in the expected manner, as the concentration of methanol increases the retention volume of the ethanol

two methyl groups next to the silicon atom hinder the reaction of adjacent hydroxyl groups with the reagent and thus there will be a considerable amount of unreacted hydroxyl groups remaining even after capping. In the extreme, it has been suggested that there is a hydroxyl group situated between each bonded chain. There is certainly evidence of some polar interactions with reverse phases which if completely covered with hydrocarbon chains should only exhibit dispersive interactions. However, reverse phases are predominantly dispersive in character and it would appear that if there are any hydroxyl groups still present on the surface it is likely that they would be relatively few in number compared with the bonded moieties

The spatial character of the solute will determine the degree of entry and consequently the proximity of interaction which, in turn, will determine the energy of interaction and the magnitude of the retention. Finally, the fifth group contains the cyclodextrin based materials that control retention in a similar manner to that previously described for GC. In LC, the cyclodextrin stationary phases are bonded to a support such as silica and are prepared using similar techniques to those for making reverse phases. The more recent and most effective stationary phases are without doubt those based on the macrocyclic glycopeptides and the cyclodextrins

commercially available reverse phases and determined the carbon content of each phase and the retention volume of a series of solutes on columns packed with each adsorbent. The retentive properties of the five reverse phase are shown in figure 37 where the corrected retention volume (V'r) of 2-ethyl anthraquinone is plotted against carbon content of the reverse phase. It is seen, somewhat surprisingly, that there is a linear relationship between retention volume and carbon content of the brush phases (R2, R8, R18). This relationship can only be expected to occur if all the stationary phase is available to the solute and the packing procedure is very reproducible so that each column contains the same amount of packing.   Figure 37. Graph of Retention Volume against Carbon Content (%w/w) It should again be stressed that all three reverse phases were produced from base silicas of very different surface areas and, despite this, the linear relationship between carbon content and

Short chain reverse phases reduce the extent to which proteins are denatured in the separation of substances of biological origin, it is seen by the chromatogram from the C2 reverse phase, that a serious price must be paid in loss of resolution if the nature of the separation demands the use of such material. However, the development of the polymer packings have, at last, partly solved this problem. In general, because the brush type phases can be synthesized in a more reproducible manner, particularly if carried out in a fluidized bed, the brush phases are generally recommended for the majority of applications. For high retentive capacity and for systems that will be operated with aqueous solvent mixtures having a very high water content, the bulk phases might be preferred. The partially reacted, low carbon content bulk phase also have special areas of application particularly in sample preparation. Macroporous

Brush Phases. The equation for the corrected retention volume of a solute (V'r ) (see Plate Theory and Extensions ) is as follows,                            V'r  =  KVS  or    V'r  =  KAS where (K) is the distribution coefficient of the solute between the two                phases,          (VS) is the effective volume of stationary phase in the column,   and  (AS) is the effective surface area of the packing. The two equations are given to illustrate that the reverse phase system may be considered as a liquid/liquid or liquid/solid distribution system where,                                          VS  =  dfAS   and (df) is the effective film thickness of the bonded material and                any  solvent that may be adsorbed on its surface