The accelerated transformation of three azithromycin pseudo-polymorphs (viz. the anhydrous, monohydrate (MH), and dihydrate (DH) forms) at a high temperature and moisture level was examined by near infrared spectroscopy (NIRS). The most marked spectral differences between the pseudo-polymorphs occurred in the 1800-2200 nm region, which corresponds to the first overtone for water. The qualitative analysis of the NIR spectra for the pseudo-polymorphs following storage in a stove at 608C at 100% relative humidity for 60 days suggests that the crystalline forms (viz. the MH and DH) are stable, whereas the amorphous (anhydrous) form evolves to the DH. This was confirmed by determining the amounts of water and DH present in anhydrous azithromycin and the MH by use of partial least-squares regression (PLSR). The method used to quantify the DH in MH samples was developed and validated in accordance with the standards of the International Conference of Harmonization (ICH) and the European Medicines Agency (EMEA) with a view to its subsequent application by the pharmaceutical industry. The limits of detection (LD) and quantitation (LQ) for the DH in MH provided by the NIRS method were consistent with those obtained by X-ray diffraction (XRD) methodology. This testifies to the accuracy of the proposed method.