A fast, sensitive, selective, and biocompatible dual sensor of an antineoplastic medication (methotrexate) and a neurotransmitter (adrenaline) is still being searched by present-day scientists. To overcome this issue, we have designed a functionalized, robust, bio-friendly luminescent MOF for the sensitive, selective, and rapid monitoring of methotrexate and adrenaline. This probe is the first ever reported MOF-based fluorescence sensor of methotrexate and second only for adrenaline. This fluorescence probe has a very low limit of detection (LOD) of 0.34 and 11.2 nM for adrenaline and methotrexate, respectively. The sensor can detect both the targeted analytes rapidly within 5 s. It can also detect adrenaline and methotrexate from human blood serum and urine accurately and precisely. This reusable sensor is equally efficient in detecting methotrexate from environmental water specimens. Biocompatible, user-friendly, and inexpensive chitosan@MOF@cotton composites were fabricated for the detection of adrenaline and methotrexate from the nanomolar to the micromolar range by the naked eye under a fluorescence lamp. This probe displayed high reproducibility, precision, and accuracy in sensing methotrexate and adrenaline. Fluorescence resonance energy transfer (FRET) and the inner filter effect (IFE) are the possible mechanisms for adrenaline and methotrexate sensing, respectively. The possible mechanism was supported by using required instrumental techniques and theoretical simulations.