A Comparative Study of the Kinetics of Cytarabine Hydrolytic Deamination in Aqueous Solutions

Abstract

Cytarabine (1-D-arabinofuranosylcytosine, I) is a drug belonging to the group of nucleoside antimetabolites [1]. At present, the drug is produced abroad both in dry (lyophilized) and liquid forms. The production of liquid forms is less labor-consuming and more economically effective. At the same time, it was reported [2 – 8] that cytarabine is subject to hydrolytic deamination in aqueous media with the formation of a therapeutically inactive 1-D-arabinosyluracil (II) and ammonia (Fig. 1). Our task was to study the stability of cytarabine in aqueous solutions during a preset storage time. For this purpose, it was necessary to select a method that would be (i) capable of studying the kinetics and rate of the cytarabine hydrolytic deamination reaction and (ii) sufficiently simple and optimum from the standpoint of sensitivity and reproducibility. Various methods were developed for determination of the content of I in medicinal preparations and for the study of hydrolytic deamination of this compound. These include TLC [9], RIA [10 – 13], mass spectrometry [14], gas chromatography in various modes, including those combined with other techniques [15, 16], UV spectrophotometry [17], extraction methods [18], and liquid chromatography at low [19] and high pressures [20 – 35]. The method of direct UV spectrophotometry [17] is based on the difference between the absorption spectra of I and its decomposition product II (having absorption peaks at 280 and 260 nm, respectively, in 0.1 N HCl). This technique is most simple and was widely used in the early investigations of the hydrolytic deamination reaction [2 – 4, 6 – 8]. In recent years, compounds I and II are more frequently determined by HPLC techniques offering some advantages over the other methods. Compounds belonging to arabinosyl nucleosides were predominantly studied by inversed-phase HPLC (IPHPLC) [20 – 22], although ion-exchange HPLC [23] and combined techniques can be used as well [27]. In application to nucleosides, IPHPLC can be used in several different modes [36], including chromatography under socratic conditions (most frequently used for the analysis of arabinosyl nucleosides) [20 – 22], gradient-elution chromatography, and ion-pair chromatography [29, 34, 35]. As is known [3], the maximum extinction coefficient of cytosine nucleosides is observed at low pH. Therefore, using these conditions we may reach a maximum sensitivity of spectrophotometric detection. At the same time, all the molecules of cytosine nucleosides studied under these conditions are ionized and, hence, exhibit weak adsorption on the immobile (nonpolar) phase that results in lower resolution. Thus, optimum possibilities for controlling the interaction of partitioned components with the immobile phase are provided by the method of ion-pair chromatography.