Recently, scientists developed a powerful strategy called “one drug-multiple targets” to discover vital and unique therapies to fight the most challenging diseases. Novel derivatives of isatin-based Schiff bases 2–7 have been synthesized by the reaction of 3-hydrazino-isatin (1) with aryl aldehydes, hetero-aryl aldehydes, and dialdehydes. The structure of the synthesized derivatives was proved by physical and spectral analysis. Additionally, in vitro biological studies were performed, including antioxidant, anti-diabetic, anti-Alzheimer, and anti-arthritic activities. The four derivatives 3b, 5a, 5b, and 5c possess the highest activities. Among the four potent derivatives, compound 5a exhibited the highest antioxidant (TAC = 68.02 ± 0.15 mg gallic acid per g; IRP = 50.39 ± 0.11) and scavenging activities (ABTS = 53.98 ± 0.12% and DPPH = 8.65 ± 0.02 μg mL⁻¹). Furthermore, compound 5a exhibited an α-amylase inhibitory percentage of 57.64 ± 0.13% near the acarbose (ACA = 69.11 ± 0.15%) and displayed inhibitor activity of the acetylcholinesterase (AChE) enzyme = 36.38 ± 0.08%. Moreover, our work extended to determining the anti-arthritic effect, and compound 5a revealed good inhibitor activities with very close values for proteinase denaturation (PDI) = 39.59 ± 0.09% and proteinase inhibition (PI) = 36.39 ± 0.08%, compared to diclofenac sodium PDI = 49.33 ± 0.11% and PI = 41.88 ± 0.09%. Additionally, the quantum chemical calculations, including HOMO, LUMO, and energy band gap were determined, and in silico ADMET properties were predicted, and their probability was recorded. Finally, molecular docking simulations were performed inside α-amylase and acetylcholinesterase enzymes.