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A simple and sensitive method that dose not require derivatization for determining cholestanol has been developed using HPLC with electrochemical detection (HPLC-ECD). The current peak height was linearly related to the amount of cholestanol injected, ranging from 1 to 200 μM (r = 0.999). The detection limit (S/N = 3) of cholestanol was 0.23 μM (1.2 pmol). Total cholestanol in control human and mouse serum was determined by the present method with a recovery rate of more than 90% and an RSD (n = 5) of less than 7.3%. Further, this method was successfully applied to monitor experimental hypercholestanolemia in mice fed a high-cholestanol diet, an animal model of cerebrotendinous xanthomatosis (CTX). In conclusion, we found this method to be both simple and useful for the determination of cholestanol in serum, helping in the diagnosis of CTX.

The aim of this study was to establish a novel approach to in vitro dissolution evaluation using a combination of the paddle method and a dialysis membrane, both to predict the overall in vivo performance of tacrolimus microspheres and also to identify a suitable dissolution test method to describe the in vivo initial burst phenomenon. This new dissolution method for evaluating the release of tacrolimus from microspheres consisted of rotating a customized paddle inside a dialysis membrane using a conventional paddle apparatus. Findings were compared with a method in which the paddle was rotated outside the dialysis membrane, the conventional paddle method, and the flow-through cell method. We concluded that the paddle method with a dialysis membrane and internal agitation, which was designed to mimic in vivo conditions, predicted the overall pharmacokinetic (PK) profile of tacrolimus microspheres whereas the conventional paddle method described the initial burst. These findings suggest that it may not be possible to predict both the PK profile and initial burst using a single analysis method. We therefore recommend that evaluation of the initial burst be performed separately. In conclusion, we propose that combination of the paddle method with a dialysis membrane and internal agitation to evaluate the overall PK profile, together with the paddle method to describe the in vivo initial burst, represents a novel approach to in vitro dissolution evaluation for microsphere formulations.