Radiation effect on nucleosides: 3. Radiolysis of cytidine

Abstract

The studies of processes caused by the action of ionizing radiation on nucleic acids and their main constituents (nitrogen bases, nucleosides, and nucle otides) have been actively performed for almost half a century. The development of analytical techniques such as HPLC-MS/MS and GC-MS made it possi ble to identify the main products formed by the action of radiation (mainly, different hydroxy compounds, which appear as a result of the addition of the OH rad ical or Н 2 О at the double bond of a nitrogen base) [1][2][3]. Cytidine remains the least studied compound in comparison with other pyrimidine nucleosides. This is explained by the lower stability of radiolysis products at the initial stage of the irradiation of cytidine. The mechanism and kinetics of the processes are unclear. In this paper, we consider the kinetics of cytidine deg radation and the buildup of the resulting products, which is important for explaining the mechanisms of the processes of interest.Cytidine (Sigma Aldrich, >99%) and cytosine (Fluka, >99%) in glass ampoules were irradiated on an RKhM-γ-20 source in a nitrogen atmosphere. Dosimetry was performed in accordance with the Fricke method. The dose rate was 62 Gy/h.The gas phase of irradiated solid cytidine was ana lyzed on a GChF 18.3 Chromatron chromatograph. A column (3 m × 4 mm) packed with activated carbon was used. Nitrogen was a carrier gas, and a katharom eter was a detector.Cytidine.The chromatographic analysis of the radiolysis products of the aqueous solutions of cytidine was per formed on an LC 10AD HPLC instrument with a UV SPD 10А VP detector (Shimadzu) at 260 nm using a C18 column and a mobile phase of acetonitrile-water (2 : 98). The mass spectra of radiolysis products were measured on an HPLC/MS Agilent Technologies 6410 Triple Quad LC/MS instrument (C18 column; gradient elution; electrospray ionization).Hydrogen is the main gas product of the radiolysis of solid cytidine. The radiation chemical yield of Н 2 was G = 0.12 ± 0.01 1/100 eV, which is somewhat lower than that upon the irradiation of thymidine [4]. The intensity of light absorption (λ = 260 nm) by the irra diated aqueous solutions of cytidine decreased with dose analogously to the behaviors of uridine and thy midine. However, the conversion of cytidine occurred more slowly than that in the cases of uridine and thy midine. The HPLC analysis made it possible to quan titatively characterize these changes. The radiation chemical yield of disappearance of cytidine at low conversions was 3.0 1/100 eV. Chromatographic anal ysis (HPLC) demonstrated that during the irradiation of the aqueous solutions of cytidine in a dose range of 0.1-1500 Gy, it transformed via a few reaction paths: cytidine decomposed to cytosine by the cleavage of the N glycosidic bond at low doses; and the addition of the OH radical at the double bond of the base subse quently led to the formation of 5 hydroxycytidine and cytidine glycol with the deamination of the latter to uridine glycol.The transformation of cytidine in an aqueous solu tion by th...