This research was motivated by the low level of environmental literacy among biology education students, as well as the limited integration of Islamic values into instruction on environmental pollution. A preliminary study indicated an initial environmental literacy score of 45%, falling into the "low" category. The objective of this study was to develop an E-Module on environmental pollution—infused with Islamic values—that is valid, practical, and effective for enhancing students' critical thinking skills and environmental literacy. The study employed the ADDIE model of Research and Development (R&D), combined with a quasi-experimental, pre-test–post-test control group design. The participants included 94 students from Universitas Al-Washliyah Labuhanbatu and Universitas Labuhanbatu. Data were collected using validation questionnaires, practicality questionnaires, and environmental literacy questionnaires. Data analysis involved descriptive statistics, prerequisite testing, ANCOVA (as the primary analysis), supplementary t-tests, and N-Gain calculations. The results demonstrate that the E-Module met the criteria for being highly valid (89.6%) and highly practical (96.25% according to lecturers; 90.25% according to students). ANCOVA results revealed significant differences in critical thinking skills (F = 25.34; p < 0.001) and environmental literacy (F = 30.12; p < 0.001) after controlling for pre-test scores. The N-Gain values for the experimental group (0.63 and 0.66) were higher than those for the control group (0.31 and 0.35). In conclusion, the E-Module, infused with Islamic values, is valid, practical, and effective in improving students' environmental literacy. This E-Module can serve as an innovative teaching resource for environmental biology courses, based on the integration of science and Islamic values.