Development of a gel permeation chromatographic assay to achieve mass balance in cellulose acetate phthalate stability studies

Abstract

Cellulose acetate phthalate (CAP, cellulose acetate 1,2-benzenedicarboxylate) is a common polymeric oral tablet coating. CAP is also a vaginal microbicide candidate that potently inhibits HIV-1 proliferation. This paper describes the development of a precise, stability-indicating gel permeation chromatography (GPC) assay for CAP. During accelerated stability studies monitored by separate reversed-phase high performance liquid chromatography (RP-HPLC) and GPC analyses, an apparent loss of mass balance was observed. This deficit was corrected by recalculating the response factor (RF) for each degraded sample, proportional to the fraction of phthalate remaining bound to the polymeric CAP. The correction factor enabled CAP and the degradation product phthalic acid (PA) to be quantitated by a single GPC analysis. The chromatographic approach taken here could potentially apply to any polymer containing degradable chromophores.

Fig. 1

Structures of cellulose acetate phthalate (CAP), phthalic acid, and phthalic anhydride.

Fig. 2

24-hour forced degradation of CAP at 100 °C assayed by GPC

Fig. 3

Gel permeation chromatography control injections are shown with no x-axis offset. The polystyrene 104 kDa size marker control ran at approximately the same retention time as CAP. The acetone control at 15.5 minutes retention time marks the lower size limit that can be resolved using the GPC method. Acetic acid was not retained by GPC. A PA standard injection is shown for comparison. Inset: Acetic acid and phthalic acid calibration standard injections as analyzed by the RP-HPLC method.

Fig. 4

Bound Phthalate (BP) calculated at each stability time point using Equation 1. Measured quantities are shown as solid lines and calculated quantities are dashed lines. Total phthalate (TP) was measured by RP-HPLC after hydrolysis, and phthalic acid (PA) by RP-HPLC was subtracted from it to give BP. PA + phthalic anhydride by GPC is closer to the RP-HPLC PA peak than PA by GPC is to PA by RP-HPLC, suggesting that PA and phthalic anhydride coelute in the RP-HPLC assay.

Fig. 5

Linearity of the CAP RF correction factor. A linear relationship between the corrected CAP RF and BP/TP is predicted by Equation 5.

Fig. 6

Normalized UV spectra acquired by the DAD of an Agilent 1100 during GPC analysis. Triangles: CAP. Squares: PA. Diamonds: Phthalic anhydride. For comparison purposes, absorbance values are shown as a fraction of the maxima of 574 mAU at 210 nm for CAP, 203 mAU at 210 nm for PA, and 59.0 nm at 212 nm for phthalic anhydride. Inset: Spectral data to scale.