Determination of reducing sugars is carried out routinely in the food industry, in biological research, or pharmaceutical and biomedical quality control to estimate metabolically assimilable sugars. Widespread detection methods are complex, expensive, or highly polluting. Here, we propose the use of spectrophotometric quantification for reducing sugars (Benedict q ) based on the qualitative method of Benedict. The protocol was validated, to verify its reproducibility and precision. With the proposed method (Benedict q ), the reducing sugar glucose can be determined in a range of 0.167–10 mg mL –1 , with an R 2 of 0.997 and accuracy (expressed as % of recovery) greater than 97%. Other reducing sugars, such as maltose, fructose, and lactose, showed similar values. The method robustness was verified for pH values greater than or equal to 4. In the case of protein presence, a correction is proposed in the range of 0–1.67 mg mL –1 . Modifications implemented in the protocol reduce cost, working time, and reaction volumes with respect to the original assay without detriments in accuracy and precision. In addition, waste reduction represents an important contribution of the method.