hilic
65
hilic
Direct connection HILIC column for dedicated applications
The TSKgel NH
2
-100 DC column connects directly to other TSKgel HPLC
columns. This can be used to combine separations based on polar in-
teractions and non-polar interactions e.g. HILIC/ion exchange and re-
versed phase without the need of connectors or capillaries. The DC in
the name ‘TSKgel NH
2
-100 DC’ emphasizes this ‘direct connect’ aspect.
A male-type outlet end fitting enables the direct connection to the nor-
mal end fitting of a TSKgel reversed phase column. This allows for the
simultaneous gradient separation of hydrophobic and hydrophilic/aci-
dic compounds - e.g. an active pharmaceutical ingredient (API) and its
counter ion - without the loss of column efficiency normally experienced
when connecting two columns with capillary tubing. Hydrophilic com-
pounds and anions are retained strongly on the amino-alkyl bonded
3 µm silica phase of the TSKgel NH
2
-100 DC 3 µm column. When coupled
to a reversed phase column the overall retention of these compounds is
thereby shifted from other unretained peaks.
Figure 9
demonstrates the use of the TSKgel NH
2
-100 DC column in the
separation of drug and counter ion. Maleic acid and p-toluene sulfonic
acid are commonly used as counter ions in pharmaceutical prepara-
tions. Both of these organic acids are hydrophilic and are not retained
on a TSKgel ODS-100V reversed phase column at pH 7.0 in 70 % me-
thanol eluent (Chromatogram B). With the connection of a TSKgel NH
2
-
100 DC column prior to the TSKgel ODS-100V column, the simultaneous
determination of maleic acid and the active pharmaceutical ingredient
(API) desipramine becomes possible (Chromatogram A). Maleic acid
is slightly retained on the TSKgel NH
2
-100 DC column by an anion ex-
change interaction. Desipramine, on the other hand, does not interact
with the protonated amino groups as it is positively charged.
figure 9
Simultaneous analysis of maleic acid and desipramine
Columns:
A: TSKgel NH
2
-100 DC 3 µm + TSKgel ODS-100V 3 µm
B: TSKgel ODS-100V 3 µm
Mobile phase: 50 mmol/L phosphate buffer, pH 7.0/MeOH (30/70); Flow rate: 1.0 mL/min; Detection: UV @ 210 nm; Temperature: 40°C; Injection vol.: 5 µL;
Samples: 1. maleic acid (50 mg/L); 2. p-toluene sulfonic acid (50 mg/L); 3. desipramine (50 mg/L)
