Hyperpolarized (HP) xenon-129 (Xe), when dissolved in blood, has two NMR resonances: one in red blood cells (RBC) and one in plasma. The impact of numerous blood components on these resonances, however, has not yet been investigated. This study evaluates the effects of elevated glucose levels on the chemical shift (CS) and T* relaxation times of HP Xe dissolved in sterile citrated sheep blood for the first time. HP Xe was mixed with sheep blood samples premixed with a stock glucose solution using a liquid–gas exchange module. Magnetic resonance spectroscopy was performed on a 3T clinical MRI scanner using a custom-built quadrature dual-tuned Xe/H coil. We observed an additional resonance for the RBCs (Xe-RBC1) for the increased glucose levels. The CS of Xe-RBC1 and Xe-plasma peaks did not change with glucose levels, while the CS of Xe-RBC2 (original RBC resonance) increased linearly at a rate of 0.015 ± 0.002 ppm/mM with glucose level. Xe-RBC1 T* values increased nonlinearly from 1.58 ± 0.24 ms to 2.67 ± 0.40 ms. As a result of the increased glucose levels in blood samples, the novel additional HP Xe dissolved phase resonance was observed in blood and attributed to the Xe bound to glycated hemoglobin (HbA).