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Assaying Local Anesthetics by GC/FID

Optimizing System Suitability, Using an Rxi-5ms Column


By Rick Lake, Pharmaceutical Innovations Chemist
  • Rxi-5ms column assures excellent peak shapes for basic compounds.
  • Stable, reproducible retention times.
  • Easy conformance to stringent system suitability criteria.

Local anesthetics are biologically active compounds that reversibly inhibit the propagation, or broadcasting, of signals along nerve cell pathways. Because of this action, they are widely used as drug compounds to produce temporary analgesia (loss of pain) and paralysis (loss of muscle movement). Anesthetic compounds are formulated into a large number and wide variety of drug products, ranging from over-the-counter topical ointments to clinical injectables, and they often are formulated in combination with other active ingredients. Therefore, many analyses of local anesthetics involve manufacturing assays, like potency and stability assays, which require high throughput and reproducible results. These assays require the fulfillment of system suitability criteria and, for this reason, we investigated assaying local anesthetics by GC/FID, using common system suitability parameters as evaluation criteria.

By GC standards, a local anesthetic is a high molecular weight, weakly basic, active compound. We took these characteristics into account when we chose the column and inlet liner for this application. Considering that these analytes are basic and active, the deactivation of the inlet liner and capillary column is very important. For superior inertness, we chose to use an Rxi-5ms column.

When analyzing high molecular weight compounds — the normal case in pharmaceutical assays — discrimination and irreproducible injections sometimes occur, primarily due to incomplete vaporization of the analytes. This can be especially problematic for analysts who must meet stringent system suitability criteria. Some liners, like the laminar cup and cup splitter, were designed specifically for samples containing high molecular weight compounds. These liner designs aid in sample vaporization, but at a cost of reduced internal volumes and intricate flow paths that can cause poor reproducibility when such liners are used with a solvent that has a large expansion volume, like methanol. In this application, we used our conventional, intermediate polarity deactivated, split liners packed with intermediate polarity deactivated wool. Wool in the liner provides a large surface area, for rapid vaporization, but the liner still delivers a uniform vapor cloud to the split point.

Under these conditions, chromatography from a six-replicate system suitability analysis (Figure 1) was well within normal acceptance criteria (Table 1). USP tailing, approximately 1.00 for all analytes, shows the exceptional inertness of the Rxi-5ms column. In addition, retention times and area responses were extremely stable. The Rxi-5ms column, coupled with an appropriate inlet liner, provides the stability and deactivation necessary to afford easier conformance to system suitability criteria.


Figure 1  An Rxi-5ms column provides excellent peak shape and stable retention times for basic compounds, for easier conformance to system suitability criteria.

Peaks
1.Benzocaine
2.Prilocaine
3.Lidocaine
4.Procaine
5.Tetracaine
6.Bupivacaine
7.Dibucaine
Local Anesthetics Rxi-5ms
GC_PH00860
ColumnRxi-5ms, 30 m, 0.53 mm ID, 1.00 µm (cat.# 13455)
Sample
Diluent:methanol
Conc.:50 µg/mL each component
Injection
Inj. Vol.:1.0 µL split (split ratio 10:1)
Liner:4mm Split w/Wool (cat.# 20781)
Inj. Temp.:250 °C
Oven
Oven Temp.:200 °C (hold 4 min) to 320 °C at 30 °C/min (hold 3 min)
Carrier GasHe, constant flow
Flow Rate:5.0 mL/min
DetectorFID @ 300 °C

Table 1  An Rxi-5ms column provides exceptionally stable retention times and area responses.

Compound
Peak Area
(%RSD)
Retention
Time (%RSD)
USP Tailing
Efficiency
benzocaine
0.85
0.03
1.00
55858
prilocaine
1.36
0.02
1.00
(isothermal)
lidocaine
1.01
0.02
1.00
 
procaine
1.83
0.03
1.00
 
tetracaine
1.78
0.01
1.00
 
bupivacaine
1.64
0.02
1.02
 
dibucaine
1.17
0.06
1.00
 
Mean
1.38
0.03
1.00
 
six-replicate system suitability analysis

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