Separation of linear gramicidins using carbon dioxide-containing mobile phases

Thurbide, Kevin V.; Jianmin Zhang
July 2005
Analytical & Bioanalytical Chemistry;Jul2005, Vol. 382 Issue 5, p1227
Academic Journal
Packed-column supercritical-fluid chromatography (pSFC) is presented as a novel method for separating and analyzing gramicidin samples. By use of methanol-modified carbon dioxide as a mobile phase the pentadecapeptides gramicidin A (gA), gramicidin B (gB), and gramicidin C (gC) are readily separated and eluted from a PRP-1 poly(styrene–divinylbenzene) column. Although optimum separation conditions are typically achieved near a column temperature of 40°C, a column pressure of 11 MPa, and 30% methanol modifier, pressure and modifier gradients around these values are also found to improve the overall separation time. Measurements indicate that the mobile phase solubility of gramicidin under these conditions is 5.0±0.4 μg mL−1. Collection of individual peaks during chromatography achieved analytical-scale isolation of 2 μg refined gC from 20 μg injected gramicidin D. Further, supercritical-fluid extraction of 200 μg gramicidin D from a Chromosorb 102 support packed into the vessel produced 57 μg gA in 90% purity. The results establish that carbon dioxide-based mobile phases can be successfully used for the separation of individual gramicidin species.


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