Agricultural byproducts such as banana peel and corn husk are cost-effective and abundant sources of lignocellulosic biomass, offering significant potential for producing value-added biological and biochemical products. Chemical composition analysis shows that banana peels contain 13–15% hemicellulose, while corn husks contain 40–50%, making them valuable sources of xylans, particularly for Xylo-oligosaccharide production. This study explored the enzymatic hydrolysis of xylans from banana peels and corn husks using xylanase concentrations ranging from 2 to 10 mg/mL at 55 °C for 30 minutes. Samples were prepared as untreated samples or autoclaved (121 °C, 15 min). Reducing sugar concentrations were positively correlated with xylanase concentration, with the highest levels observed at 10 mg/mL. For banana peels, the maximum reducing sugar concentrations were 3.47 mg/mL for untreated samples and 3.93 mg/mL for autoclaved samples, while corn husks yielded 3.35 mg/mL and 2.88 mg/mL, respectively. Notably, banana peels exhibited higher reducing sugar values compared to corn husks, and autoclaved samples consistently outperformed untreated ones within each biomass type. Thin-layer chromatography (TLC) confirmed that the hydrolysis products of both untreated and autoclaved samples from banana peels and corn husks consisted of Xylo-oligosaccharides, including xylobiose through xylohexaose. The presence of these oligosaccharides highlights the efficiency of enzymatic hydrolysis in breaking down xylans into functional biomolecules. These findings emphasize the potential of banana peels and corn husks as sustainable and economical raw materials for Xylo-oligosaccharide production, contributing to the valorization of agricultural waste. This approach aligns with the principles of green biotechnology, supporting the development of renewable resources and advancing waste-to-value strategies for biotechnological applications.