Neutral Bonded Phases

Diol silica phases were among the first bonded silica gek to be developed in order to overcome the disturbing adsorption properties of underivatized siUca. Chemically bonded diol phases exhibit high polarity and hydrogen bonding 

A summary of the most common products and their structural and technical properties is given in Table 7.2. 

Brand name Manufacturer Particle size (pm) Pore size (A) Surface area (m /g) 

Cyanopropyl-bonded silica phases emanate from the early days of normal-phase liquid chromatography, although they can also be used in the RP mode. 

Due to the absence of hydrogen donor capabilities [31], cyanopropyl silica phases are less retentive in normal-phase liquid chromatography than under-ivatized silica or other NP packing materials. Therefore, very few applications have been reported that utilize cyanopropyl-bonded silica in the HILIC mode [32,33]. The limited number of applications may also be attributed to the mechanical instabiUty of cyanopropyl-bonded silica when operated with solvents of intermediate polarity. This instabihty is caused by a decrease in the adhesion of particles to each other that maintain the integrity of the column bed in either nonpolar or highly polar solvents [25]. Dinh et al. [34] performed a multivariate modeling of column selectivity by principal component analysis of chromatographic data from polar compounds of various structures on 20 commercially available HILIC columns and verified the low potential of cyanopropyl-bonded silica columns due to insufficient hydrophilicity. 

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These cements are formed by a similar process to the silicate minerals described in Chapter 1, the difference being the rate. Silicate minerals are formed at a rate lower by orders of magnitude compared with dental cements. In the case of dental cements, the phosphoric acid releases protons in the solution and lowers its pH. This decomposes the glass and releases silicon in the solution, and silicic acid forms as an intermediate product [26,27]. Simultaneously, cations such as Ap, Ca " ", and Na" " and the anion F are also released [28]. The cations and anions are attracted to each other, and neutral bonding phases form. Such a bonding network, especially that of aluminum, results in gelation and subsequent polymerization of a hard product. 

The stationary phase of MIC supports consists of metal ions coordinated by chelating ligates affixed to a neutral bonded phase for HPLC, the matrix must also be rigid. Figure 2 illustrates the hypothetical surface of an HPLC stationary phase for MIC of proteins. The stationary phases for high-performance MIC of biopolymers must meet the following criteria ... 

Scientific (Northbrook, IL) contain a silica support with a -y-glycidoxypropylsi-lane-bonded phase to minimize interaction with anionic and neutral polymers. The columns come in five different pore sizes ranging from 100 to 4000 A. The packing material has a diameter from 5 to 10 /cm and yields in excess of 10,000 plate counts. With a rigid silica packing material, the columns can withstand high pressure (maximum of 3000 psi) and can be used under a variety of salt and/or buffered conditions. A mobile phase above pH 8, however, will dissolve the silica support of the column (21). A summary of the experimental conditions used for Synchropak columns is described in Table 20.8. 

One disadvantage of all silica-based stationary phases is their instability against hydrolysis. At neutral pH and room temperature the saturation concentration of silicate in water amounts to lOOppm. Solubility increases with surface area, decreasing particle diameter drastically with pH above 7.5. This leads also to a reduction of the carbon content. Hydrolysis can be recognized during the use of columns by a loss in efficiency and/or loss of retention. Bulky silanes [32], polymer coating [33], or polymeric encapsulation [34] have been used in the preparation of bonded phases to reduce hydrolytic instability, but most of the RPs in use are prepared in the classical way, by surface silanization. Figure 2.3 schematically shows these different types of stationary phases. 

Table 3 Selection of Commonly Used Neutral Polar-Phase Modifying Reagents, Which at pH <7.0 Modify the Retention Characteristics of Unprotected Peptides on Chemically Bonded Hydrocarbonaceous Stationary Phasesab...

Ozretich RJ, Schroeder WP. 1986. Determination of selected neutral priority pollutants in marine sediment, tissue, and reference materials utilizing bonded-phase sorbents. Anal. Chem. 58 2041-2048. 

Another stability that should be considered in the choice of solid adsorbents is their ability to tolerate a wide pH range without loss of accumulation efficiency. All of the neutral synthetic polymers are unaffected by extremes in pH such is not the case with the bonded phases whose adsorption properties are stable only with pH values near 7. 

Kim and Salem (33) found that the acidic PL were eluted from an aminopropyl bonded phase with over 95% recovery with 4 ml of a mixture of H/2-P/Ethanol/0.1 M aqueous NH4-ac-etate/formic acid (420/350/100/50/0.5) containing 5% phosphoric acid (Table 2). Neither the solvent mixture without phosphoric acid nor methanol containing 5% phosphoric acid was able to elute the acidic PL. Actually, Table 2 indicates that a fractionation of both neutral and acidic PL is enabled by the sequential elution with methanol and the aforementioned solvent mixture. In order to remove the phosphoric acid from the acidic PL fraction, it is first dried under N2 for 10 min to remove the hexane and then extracted three times with 1 ml of chloroform after the addition of 1 ml of water. 

In fact, the results concerning J-CD activity in mobile phase solutions, concerning positional isomers of disubstituted benzene derivatives (23,28) seem to be consistent with those obtained by other authors using f-CD silica bonded phases (4-8). It should be mentioned that all of the above considerations concern mainly neutral molecules for the compounds undergoing dissociation, the equilibria are more complicated and the sequence of elution of isomers more variable. 

L. L. M. Glavina and F. F. Cantwell, Origin of indirect detection in the liquid chromatography of a neutral sample with an ionic probe using an ODS bonded phase and aqueous mobile phase. Anal. Chem. 65 (1993), 268-276. 

The type of pH modifier to make a desired mobile phase pH also has an effect on the column stability, and this is indirectly related to the peak efficiency and the retention of the analyte. As an increasing number of column volumes of the mobile phase are traversed through the column, the stability of the packing material could be comprised. Rearrangement of the packing bead leads to the loss of efficiency, dissolution of silica leads to loss in efficiency and retention, and hydrolytic decomposition of the bonded phase could impact the peak shape and retention. Different compounds, such as neutral compounds, acidic compounds, and basic compounds, could show different behaviors. 

Capillaries bonded with polymeric coatings are also applied to capillary electrochromatography (CEC) for separation of neutral molecules. In this case, the polymeric coating participate to solute-bonded phases interaction in a manner similar to open-tubular LC. Most polymeric coating preparations have followed the procedures typically used in open-tubular LC and GC. 

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