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How do intrinsically base-deactivated phases work?

By Rick Lake, Pharmaceutical Innovations Chemist

Analyzing basic compounds can be somewhat troublesome on traditional alkyl stationary phases, namely conventional C18 columns. This is largely due to the interaction of analyte molecules with silanol groups present on the silica surface. To better understand the workings of silanol interactions, it is important to consider the composition of the support material. Silica is the most commonly used support in the production of HPLC columns, mainly because it is well-suited to high-pressure chromatographic separations, giving high efficiencies and good reproducibility. Silica offers bed and pressure stability and is highly porous, which ultimately gives rise to its large surface area, increased bonding capacity and high peak efficiencies. Silica also possesses widely-studied and effective bonding chemistries, making possible diverse analyte selectivities through a wide variety of bonded stationary phases.

If we consider, however, the chemical structure of the silica surface, it is very hydrophilic, acidic, and structurally, is comprised of various forms of silanol groups (Si-OH groups). It is not possible to effectively bond, or attach a stationary phase, to the entire silica surface. Ultimately, it is the surface of the silica particle, or the free silanol groups that causes a majority of the tailing effect of basic compounds. Basic analytes often exhibit tailing on these hydrophobic, reversed phase packings. This is mainly due to the interaction of the protonated form of the base with silanol groups on the silica surface. As this occurs, the molecules undergoing these brief interactions lag behind the main peak band, causing an elongated distribution on the latter half of the peak, or a “tail”.

For this reason, column manufacturers have created techniques for limiting the interaction of basic analytes with the surface silica. One very effective means of limiting silanol activity is by bonding a polar group, within, or intrinsic to, the hydrocarbon bonded phase. This bonding chemistry, called intrinsically base deactivated, or IBD, employs either an electrostatic barrier or polar shielding to prevent analytes from interacting with surface silanol groups. The end result is an alkyl stationary phase specifically designed to create optimum peak shape for basic compounds (Figure 1).

Figure 1  Ultra IBD offers more flexibility in method development giving excellent peak shape for highly basic compounds — even without mobile phase modifiers and regardless of mobile phase pH.

Mobile phase pH 3 USP tailing 1.10
Peaks
1.Amitriptyline
Amitriptyline on Ultra IBD, pH 3
LC_PH0443
ColumnUltra IBD (cat.# 9175565)
Dimensions:150 mm x 4.6 mm ID
Particle Size:5 µm
Pore Size:100 Å
Temp.:ambient
Sample
Diluent:mobile phase
Conc.:~100 µg/mL
Inj. Vol.:10 µL
Mobile Phase20mM potassium phosphate (pH 3):methanol (10:90)
Flow:1.0 mL/min
DetectorUV/Vis @ 254 nm


Mobile phase pH 7 USP tailing 1.13
Peaks
1.Amitriptyline
Amitriptyline on Ultra IBD, pH 7
LC_PH0444
ColumnUltra IBD (cat.# 9175565)
Dimensions:150 mm x 4.6 mm ID
Particle Size:5 µm
Pore Size:100 Å
Temp.:ambient
Sample
Diluent:mobile phase
Conc.:~100 µg/mL
Inj. Vol.:10 µL
Mobile Phase20mM potassium phosphate (pH 7):methanol (10:90)
Flow:1.0 mL/min
DetectorUV/Vis @ 254 nm

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