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We currently reporting a simple and first label free optical insulin biosensor using a novel cum naturally existing Chromene mimic receptors via one pot greener methodology. Among synthesized 4H- Chromene derivatives, 2-amino-5,7-dihydroxy-4-(4-nitrophenyl)-4H-chromene-3-carbonitrile (Compound 1c) is showed a very good colorimetric response for insulin within the linear range of 20 fM -500 pM along with theoretically measured LOD of 15.38 fM based on the increment in absorbance. Meanwhile 2-amino-5, 7-dihydroxy-4-(pyren-2-yl)-4H-chromene-3-carbonitrile (Compound 1a) is depicted an excellent quenching in fluorescence response for insulin with LOD of 7.07 fM and along with wide working range from 10 fM -600 pM. The developed biosensor is tested for the detection of insulin in human blood serum using direct as well as standard addition method and the recovery results are showed a very good agreement with standard clinical chemiluminescent assay method. Hence, this developed biosensor is more reliable for clinical detection of Insulin and so a good analytical method for Diabetes diagnosis within a short period of time., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022 Elsevier B.V. All rights reserved.)