Deanna Hurum, Brian De Borba and Jeff Rohrer, Dionex Corporation, Sunnyvale, California, USA.
Cephalosporins contain a four-member β-lactam ring that is inherently strained and prone to hydrolysis and photolysis, limiting
stability and leading to degradation of products that may be toxic.1 In addition, synthetic byproducts are generated and persist during production of these antibiotics including cefepime. Analysis
of cefepime purity is particularly challenging because of the presence of such isomeric synthetic impurities. The Acclaim
120 C18, 3 μm can be used to meet and exceed the criteria set by the USP for determining related substances and assaying the purity of cefepime.
Experimental
Figure 1
|
A Dionex UltiMate 3000 system with a DGP-3600M pump, an FLM-3100 flow manager (a TCC-3200 column compartment could be used)
a WPS-3000T autosampler and a VWD-3400 detector was used in this work. An Acclaim 120 C18, 3 μm Analytical (2.1 × 150 mm)
was used for all separations with a flow-rate of 0.20 mL/min. Analytes were detected by UV absorption at 254 nm. A 1 mL injection
volume was used throughout. Chromeleon chromatography management software (Dionex) was used for system control and data processing.
Results and Discussion
Table 1
|
Because of the smaller column dimensions and the 3 μm particle size, the USP gradient method can be considerably shortened and still meet the USP requirements. Figure 1 illustrates the separation possible with a gradient method that removes 20 min from the run time for
each injection. This shortened gradient meets the USP criteria for peak asymmetry, resolution between cefepime and cefepime related substances A and B, theoretical plates and
capacity factor. The precision of this shortened gradient was tested with retention time RSDs of 0.06% or less and peak area
RSDs of 0.82% or less (Table 1). Resolution is significantly improved, including the clear separation of an additional peak
with a retention time of 9.39 min. This peak is not described in the current USP monograph, but it is recognized as impurity F in EP method 2126.
Conclusion
In this study, the Acclaim 120 C18 column, an HPLC L1 column with good steric selectivity, was successfully used for the determination
of cefepime and cefepime related substances. The methods reduced the time needed for analysis by 40% and improved the resolution
as compared to the current USP method. Additionally, the low flow-rates used in this method save time and resources spent on mobile phase preparation and
reduce waste production.
Reference
1. S.M. Rabouan-Guyon et al., Int. J. Pharm., 154, 185–190 (1997).
Acclaim, Chromeleon, and UltiMate are registered trademarks of Dionex Corporation.
Dionex Corporation
1228 Titan Way, PO Box 3603, Sunnyvale, California 94088-3603, USA
te. +1 408 737 0700 fax +1 408 730 9403
Website: http://www.dionex.com/
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