Abnormally shaped red blood cells (RBCs), called poikilocytes, can cause anemia. At present, the biochemical abnormalities in poikilocytes are not well understood. Normal RBCs and poikilocytes were analyzed using whole-blood and single-cell methods. Poikilocytes were induced in rat blood by intragastrically administering titanium dioxide (TiO₂) nanoparticles. Complete blood count and inductively coupled plasma mass spectrometry analyses were performed on whole-blood to measure average RBC morphology, blood hemoglobin (HGB), iron content, and other blood parameters. Follow-up confocal Raman spectroscopy was performed on single RBCs to analyze cell-type-specific HGB content. Two types of poikilocytes, acanthocytes and echinocytes, were observed in TiO₂ blood samples, along with normal RBCs. Acanthocytes (diameter 7.7 ± 0.5 μm) and echinocytes (7.6 ± 0.6 μm) were microscopically larger (p < 0.05) than normal RBCs (6.6 ± 0.4 μm) found in control blood samples (no TiO₂ administration). Similarly, mean corpuscular volume was higher (p < 0.05) in TiO₂ whole-blood (70.70 ± 1.97 fl) than in control whole-blood (67.42 ± 2.03 fl). Poikilocytes also had higher HGB content. Mean corpuscular hemoglobin was higher (p < 0.05) in TiO₂ whole-blood (21.84 ± 0.75 pg) than in control whole-blood (20.8 ± 0.32 pg). Iron content was higher (p < 0.001) in TiO₂ whole-blood (697.0 ± 24.5 mg / l) than in control whole-blood (503.4 ± 38.5 mg / l), which supports elevated HGB as iron is found in HGB. HGB-associated Raman bands at 1637, 1585, and 1372 cm^{- 1} had higher (p < 0.001) amplitudes in acanthocytes and echinocytes than in RBCs from control blood and normal RBCs from TiO₂ blood. Further, the 1585-cm^{- 1} band had a lower (p < 0.05) amplitude in normal RBCs from TiO₂ versus control RBCs. This represents biochemical abnormalities in normal appearing RBCs. Overall, poikilocytes, especially acanthocytes, have elevated HGB.

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