A Simple, Rapid Atmospheric Pressure Chemical Ionization Liquid Chromatography Tandem Mass Spectrometry Method for the Determination of 25‐Hydroxyvitamin D2 and D3

Vitamin D plays a vital role not only in bone health but also in pathophysiology of many other body functions. In recent years, there has been significant increase in testing of 25‐hydroxyvitamin D (25‐ OH vitamin D), a marker of vitamin D deficiency. The most commonly used methods for the measurement of 25‐ OH vitamin D are immunoassays and liquid chromatography tandem mass spectrometry ( LC ‐ MS ‐ MS ). Since immunoassays suffer from inaccuracies and interferences, LC ‐ MS ‐ MS is a preferred method. In LC ‐ MS ‐ MS methods, 25‐ OH vitamin D is extracted from serum or plasma by solid‐phase or liquid‐phase extraction. Because these extraction methods are time consuming, we developed an easy method that uses simple protein precipitation followed by injection of the supernatant to LC ‐ MS ‐ MS . Several mass‐to‐charge ( m / z ) ratio transitions, including commonly used transitions based on water loss, were evaluated and several tube types were tested. The optimal transitions for 25‐ OH vitamin D2 and D3 were 395.5 > 269.5 and 383.4 > 257.3, respectively. The reportable range of the method was 1–100 ng/mL, and repeatability (within‐run) and within‐laboratory imprecision were <4% and <6%, respectively. The method agreed well with the solid‐phase extraction methods. J. Clin. Lab. Anal. 26:349‐357, 2012. © 2012 Wiley Periodicals, Inc.