The algal photosynthetic inhibition method for biological toxicity detection offers advantages such as rapid response and simple measurement. To address the insufficient response sensitivity of existing photosynthetic fluorescence parameters to typical photosystem I (PSI) inhibitors, this study developed a comprehensive parameter index (CPI) for aquatic biological toxicity, which is established based on the segmented inhibition characteristics of the fluorescence kinetic curve. CPI was systematically compared with the commonly used chlorophyll fluorescence parameters: the maximum quantum yield of photosystem II (Fv/Fm) and the performance index on an absorption basis (PIabs). The comparison focused on three key aspects: toxicity response time, response sensitivity, and stability, and the results indicated that, unlike Fv/Fm, both CPI and PIabs exhibited shorter toxicity response times and higher response sensitivities when exposed to typical photosystem I(PSI) and photosystem II(PSII) inhibitors. Consequently, subsequent investigations focused exclusively on evaluating the representational efficacy of these two parameters (CPI and PIabs). For the PSII inhibitor Atrazine, when CPI and PIabs were employed as response parameters, the limits of detection (LODs) for short-term inhibition (15 min) were 6.65 and 4.49 μg/L, respectively, and for long-term inhibition (4 h) they were 2.22 and 2.44 μg/L, respectively. No significant difference in response sensitivity was observed between CPI and PIabs against the PSII inhibitor; however, with respect to the PSI inhibitor Paraquat, when using CPI and PIabs for toxicity characterization the LODs were 0.68 and 1.21 mg/L for short-term inhibition (15 min), and 0.22 and 0.42 mg/L for long-term inhibition (8 h). Specifically, for PSI inhibitors, the LODs of CPI for both short-term and long-term inhibition were approximately reduced by 50% lower than those obtained with PIabs. In terms of the stability of toxicity response, the relative standard deviations (RSDs) for the PSII inhibitor Atrazine were 5.15% (CPI) and 14.14% (PIabs) after a short exposure duration of 15 min, and 4.43% (CPI) and 5.38% (PIabs) after an extended exposure duration of 4 hours. For the PSI inhibitor Paraquat, the RSDs were 10.18% (CPI) and 32.79% (PIabs) after 15 min, and 3.30% (CPI) and 13.54% (PIabs) after 8 h. These results confirmed that CPI demonstrated superior performance in ensuring the stability of toxicity characterization for both representative inhibitors. Collectively, these findings demonstrate that the CPI can be effectively applied for the highly sensitive detection of biological toxicity induced by both PSI and PSII inhibitors. This not only resolves the issue of low response sensitivity of the algal photosynthetic inhibition method toward PSI inhibitors, but also provides a critical parameter support for the unification of toxicity response indicators corresponding to PSI and PSII inhibitors.