The rapid synthesis techniques and interesting multidisciplinary applications make carbon nanodots (CNDs) stand out from semiconductor quantum dots. Moreover, CNDs derived from green precursors have gained more importance beyond chemically derived CNDs due to sustainable synthesis opportunities. However, the presence of molecular impurities or intermediates or fluorophores was neglected during the entire process. Herein, we illustrate the sustainable synthesis of CNDs from Hemigraphis alternata plant leaves with extended carbonization procedure (3 and 9 min) along with simultaneous ethylene glycol and diethyl ether solvent treatment method for the successful removal of interfering fluorophores. To unravel the distinction between purified CNDs (P-CNDs) and organic fluorescent carbon nanostructures (org-FCNs), we carried out photophysical, structural, and morphological studies. A quantum yield (QY) of 69 and 42% was observed for crude org-FCNs, and crude P-CNDs; however after purification, QY of 1% and absence of one component from the fluorescent decays curve suggest the removal of fluorophores. Further, HR-TEM and DLS studies showed the quasi-spherical amorphous particles having < 10 nm particle size for P-CNDs. Besides, in vitro biocompatibility investigation and cellular uptake assay (1–100 μg/mL) against the MDA-MB 468 cell lines proves the ≥ 95% cell viability and good internalization for both org-FCNs and P-CNDs. Hence, our study shows the presence of fluorophore impurities in plant-derived CNDs, the removal and resemblance in biocompatibility properties. Hence, this information can be considered during the synthesis and isolation of CNDs. Simple and effective removal of impurities to harvest pure carbon nanodots (CNDs) through solvent-based selective separation method, and revelation of the cocktail flourphores similar to biocompatible blue fluorescent CNDs were studied.