<p>The measurement of methemoglobin is critical for diagnosing toxic exposures and for monitoring treatment efficacy. Although spectrophotometric methods are widely used in clinical laboratories, the absence of commercially available quality control standards limits their reliability. This study aimed to validate a low-cost protocol for preparing internal quality control standards, utilizing an adjusted Evelyn–Malloy method with reduced blood dilution. NaNO₂-treated blood samples were used to generate methemoglobin at concentrations of up to 93%. Three primary levels (L2, L4, and L6) were prepared by spiking 1 mL of healthy donor blood with 50 µL of NaNO₂ at concentrations of 0.31, 1.24, and 7.3 g/L, respectively. Two additional levels (L7 and L8) were derived by further enriching L2 and L4 with 0.31 g/L NaNO₂ to investigate the additive properties of methemoglobin formation. Method validation was performed according to ICH guidelines. Levey–Jennings charts and Westgard rules were used to assess performance. Methemoglobin levels ranging from 1.12% to 55% showed stable absorbance over three days. Precision was acceptable (SD: 0.55–6.15%), with recoveries between 99 and 113%. The limits of detection and quantification were 0.04% and 0.12%, respectively. The average methemoglobin values for the quality control standards at L2, L4, and L6 were 3.32% ± 0.55, 10.38% ± 0.85, and 53.77% ± 6.15, respectively. L7 and L8 showed a consistent increase of 3%, confirming the additive nature of methemoglobin generation. This validated protocol offers a reliable, reproducible, and economical approach to quality control for spectrophotometric methemoglobin assays, enhancing their applicability in routine clinical use.</p>