Eco-benign spectrofluorimetric analysis of icariin in a nano-formulation using green quantum dots: insights from <i>in vitro</i> anticoagulant and docking studies

The analysis of nutraceuticals is crucial for assessing their safety, efficacy, and bioavailability and helps in their effective integration into health practices. Icariin (ICA), a natural flavonoid, was analyzed <i>via</i> a selective and sensitive method using a microwave-synthesized turn-off fluorescence probe. Full characterization of the probe was performed using several techniques. It was observed that fluorescence quenching was directly proportional to the increase in ICA concentration over a range of 1-20 μg mL^-1 with a linearity correlation coefficient of 0.9998. Additionally, the estimated values for the limits of detection and quantitation were 0.24 and 0.73 μg mL^-1, respectively. Growing interest in ICA's potential to prevent and/or treat diseases with fewer side effects and better bioavailability has highlighted the need for innovative new nano-formulations. ICA was encapsulated in a milk-based nano-formulation and tested <i>in vitro</i> for its anticoagulant activity, employing both extrinsic and intrinsic coagulation mechanisms. Additionally, <i>in vitro</i> blood hemolysis was accomplished to certify its suitability for further <i>in vivo</i> application. The outcomes of the <i>in vitro</i> anticoagulant assays were affirmed by a molecular docking study of ICA with different coagulation factors. Finally, the trio color-coded assessment of the method using versatile tools was implemented and juxtaposed with other reported analytical techniques for the assay of ICA. Additionally, the devised method's compatibility with the Sustainable Development Goals (SDGs) was demonstrated. This work offers a straightforward, cost-effective, sensitive methodology for the assay of ICA in nano-formulations. It gives insights into the crucial role of ICA as a prophylactic anticoagulant nutraceutical.