Nanoflowers (NFs) are flower-shaped nanoparticulate systems with a higher surface-to-volume ratio and good surface adsorption. Jaundice indicates yellow discoloration of skin, sclera, and mucus membrane and is a clinical indication of bilirubin accumulation in the blood which occurs as a consequence of the incapability of the liver to excrete bilirubin in the biliary tree or conjugate bilirubin and higher production of bilirubin in the body. Several methods have been developed so far for bilirubin estimation in jaundice like the spectrophotometric method, chemiluminescence method, etc., but biosensing methods provide advantages over traditional methods concerning the surface area, adsorption, particle size, and functional characteristics. The primary objective of the present research project was to formulate and examine the adsorbent nanoflowers-based biosensor for accurate, precise, and sensitive detection of bilirubin in jaundice. The particle size of adsorbent nanoflowers was found to be in the range of 300-600nm with the surface charge (zeta potential) in the range of -1.12 to -15.42 mV. Transmission electron microscopy and scanning electron microscopy images confirmed the flower-like morphological structure of adsorbent NFs. The adsorption efficiency of NFs for bilirubin adsorption was maximum at 94.13%. Comparative studies of bilirubin estimation in the pathological sample with adsorbent NFs and diagnostic kit displayed bilirubin concentration to be 1.0 mg/dL in adsorbent nanoflowers and 1.1 mg/dL with diagnostic kit indicating effective detection of bilirubin with adsorbent NFs. The nanoflower-based biosensor acts as a smart approach to elevate adsorption efficiency on the surface of nanoflower due to a higher surface-to-volume (SV) ratio. Graphical Abstract.