The composition of the mobile phase is determined by the requirem

The composition of the mobile phase is determined by the requirements for optimal next activity and stability of the used enzyme systems: essentially an aqueous phosphate buffer at pH 6.5-8.5 [28-31]. The conversion of acetylcholine to hydrogen selleck chemical MEK162 peroxide is most efficient at pH 8.0-8.5 [29, 30]. However, the optimal pH for the subsequent detection of hydrogen peroxide on the enzyme modified electrode has not been determined. The cation-exchange setup provides good chromatographic stability under these conditions, but a limited resolution between Inhibitors,Modulators,Libraries acetylcholine and choline [12, 29, 30]. Microdialysis samples typically contain a high concentration of choline, which may interfere with the acetylcholine signal.

A precolumn choline oxidase and catalase reactor was developed to eliminate choline from the microdialysis sample matrix [13, 21].

The ion-pair setup offers a superior resolution Inhibitors,Modulators,Libraries between acetylcholine Inhibitors,Modulators,Libraries and choline and does not Inhibitors,Modulators,Libraries require the preliminary elimination Inhibitors,Modulators,Libraries of choline [23, Inhibitors,Modulators,Libraries 31]. However, the stability of silica-based chromatographic columns is highly dependent on the mobile phase pH [32]. Inhibitors,Modulators,Libraries The feasibility of acetylcholine determination Inhibitors,Modulators,Libraries in an ion-pair chromatographic setup with amperometric detection was previously reported at pH 6.5, but the implications Dacomitinib of lowering the mobile phase pH for the sensitivity and long-term enzymatic and chromatographic stability were not investigated [23].

In the present study an ion-pair liquid chromatography method with amperometric detection was optimized and validated for acetylcholine determination in microdialysis Brefeldin_A samples.

Different mobile phase conditions were compared to improve the sensitivity, long-term enzymatic and chromatographic stability. The polymer-coated octadecyl silica column type used in this study allowed a reliable separation of acetylcholine from choline and other matrix Tanespimycin components over 4 months of intensive use.2.?Results and Discussion2.1. Method optimizationThe chromatographic parameters were chosen to obtain an optimal equilibrium between sensitivity and chromatographic stability. A microbore polymer-coated silica column with high endcapping and low octadecyl binding-density was selected to allow chromatographic stability when using a purely aqueous phosphate buffer as the mobile phase.

Typically, mobile phases with pH 8.0-8.5 have been used for detection of acetylcholine by liquid chromatography with amperometric detection [10-15]. Nevertheless, it has selleck products been demonstrated that it is feasible to detect acetylcholine in microdialysis samples when working at pH 6.5 [23]. In the present setup, no difference in sensitivity was observed within the pH range 6.5-8.5 (Figure 2A).Figure 2.(A) Normalized response for acetylcholine (10 nM) as a function of mobile phase pH.

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