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Mobile phase type CHIROBIOTIC V2, T, R, TAG Polar Ionic Mode Reversed Phase
Polar Organic Mode Normal Phase
30/70, EtOH/Heptane Table 1. Suggested method development screening mobile phases
100/0.1/0.1 (v/v/v), MeOH/HOAc/TEA 30/70, ACN/20mM NH4Ac, pH 4.0 100% EtOH
Chiral LC-MS-MS Since LC-MS has the ability to separate in the mass/charge dimension, it should be possible to screen for a chiralmethod formany analytes simultaneously. CHIROBIOTIC stationary phases operate best in both reversed-phase and polar ionicmodes, both of which are highly amenable to LC-MS. In a recent LC-MS study in our laboratory the polar ionicmode was used to test the efficacy of batch screening to identify unique selectivity, using a set of 14 basic probes differing over a wide range of pKa values, hydrophobicity and molecular weight. The extracted ion current of
Figure
1.Method development screening for Fluoxetine: CHIROBIOTIC V2 in both RP and Polar IonicMode show selectivity.
the β-blockermetoprolol was used tomonitor impact ofmobile phase variables on selectivity and retention, comparing single injection results against the probemix in order to investigate the potential for batch injections inmethod development. In one such comparison, a slight variation in enantiomer response due to ion-suppression by co-eluting peaks was observed; however, retention and selectivity were not compromised when using the probe testmix approach (Table 2). Since this approach is being developed for qualitativemethod development purposes only, this indicates that batch injections would not result in a false negative result. The results frombatch screens on the Chirobiotic TAG showed high selectivity and retention formany bases. Figure 2 shows the results fromthe amphetamines in the probemix, and these columns also showed good selectivity towards the beta-blockers and overall greater retention for bases compared to the other Chirobiotic phases.
Instrument: Waters/Micromass ZQ, SingleQuadrupole, Waters Alliance 2690
Column: Chirobiotic T, 150 cmx 4.6mm, 5 µm
Temperature: 35°C Figure 2. Partial results of batch screening on CHIROBIOTIC TAG in the Polar IonicMode (Extracted Ion Current)
Mobile Phase: 0.1%,w/v ammoniumacetate in methanol (Polar IonicMode)
FlowRate: 1mL/min
Detection: ESI, Positive IonMode, scan rangem/z 150–500
Inj. Volume: 5 µL Table 2: Conditions for simultaneous LC-MS chiral screen Figure 3. Van Deemter curve for Chirobiotic T in Polar IonicMobile Phase
The probemix was also used to study the impact of buffer (salt) type, buffer concentration, and acid/base ratio on retention and selectivity in the polar ionic mode2 Frequently, when optimising the method forMS detection or prep HPLC, a volatile salt such as ammoniumacetate or ammoniumformate replaces the traditional
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