Xylem exudate was collected from pruned spurs of <i>Vitis rotundifolia</i> Michx. Noble and <i>Vitis</i> hybrid Suwannee on three dates during late winter and spring in order to determine the influence of pruning date on the chemical profile of xylem fluid. Nineteen amino acids, seven organic acids, three sugars, NH<sub>4</sub><sup>+</sup>, NO<sub>3</sub><sup>-</sup>, and macro- and microelement concentrations were quantified. Water flux varied greatly with pruning date, but osmolarity remained fairly constant (Noble, 8.6 to 11.2 m<i>M</i>; Suwannee, 11.2 to 13.7 m<i>M</i>). Total osmolarity coincided closely to the sum of all organic and inorganic entities quantified, indicating that the major chemical components in the xylem exudate have been identified. The within-vine variance in elemental composition at a given time was often as great as between-vine or between-cultivar. Total inorganic ion concentration varied from 3.1 m<i>M</i> to 4.7 m<i>M</i>, amino acids from 3.3 m<i>M</i> to 5.1 m<i>M</i>, organic acids from 2.0 m<i>M</i> to 6.4 m<i>M</i>, and sugars from 10 µ<i>M</i> to 76 µ<i>M</i>. Glutamine was the predominant N-carrier in xylem exudate of both <i>Vitis</i> species. Amino acid concentration increased, and organic acids concentration decreased in Noble exudate later in the season, but inorganic ion and sugar concentrations varied asynchronously over time. The entire increase in amino acid concentration was a result of an increase in glutamine concentration, whereas the decline in organic acid concentration was mainly a function of reduced malic and succinic acid concentrations. Thus, although xylem fluid osmolarity remained relatively constant over time, substantial differences in chemical profile occurred as a function of pruning date.