Dramatic reductions of viral load and increased survival have been achieved in patients infected with the Human Immunodeficiency Virus (HIV) with the introduction of combination antiretroviral therapy. Currently 11 agents including nucleoside reverse transcriptase inhibitors (RTI), non-nucleoside RTI and protease inhibitors are available for the use for treatment of HIV infection. Recent studies have demonstrated that certain combinations of these drugs are advantageous over their individual use as monotherapy with an even more sustained viral suppression. Much emphasis has therefore been put on studies evaluating the interactions of these different compounds. Especially the intracellular metabolism of nucleoside RTI has been evaluated to some extent, by both in vitro and in vivo studies. These compounds need to undergo phosphorylation to their active 5'-triphoshates involving several enzymatic steps and the nucleoside concentration in the plasma may not correlate with intracellular concentrations of active drug. It is therefore of great importance to study these drugs at an intracellular level in order to evaluate their efficacy. This review summarizes the intracellular phosphorylation of Zidovudine and other nucleoside analogs investigated by in vitro experiments and the efforts of measuring the active anabolites in vivo in cells isolated from HIV infected patients on nucleoside therapy.