Mlanding tempeh is an indigenous fermented food from lamtoro (Leucaena leucocephala) seed that uses usar as the inoculum in its production. Lamtoro seed contains antinutritional compounds namely phytic acid and tannin, and toxin compounds namely mimosine. Antinutritional compounds decrease the quality of nutrients in mlanding tempeh. The purpose of the research was to investigate the changes in nutritional compounds, antinutritional and toxin compounds, total phenols, antioxidants activity, dietary fiber and dynamics population of microorganisms in pre-fermentation treatment in lamtoro seed during mlanding tempeh’s production. The analysis was carried out at the pre-fermentation stage, namely before the inoculation process. The result showed the change of nutrient compounds, antinutritional and toxin compounds, total phenols, antioxidants activity, dietary fiber, and dynamics population of microorganisms in prefermentation treatment in lamtoro seed during mlanding tempeh’s production. Moisture, protein, and fat content increased, whereas ash, carbohydrate, phytic acid, tannin, and mimosine decreased. The total phenols content, antioxidant activity, and dietary fiber content of lamtoro seed also decreased. Meanwhile, the population of microorganisms (yeast and lactic acid bacteria) increased in pre-fermentation treatment in lamtoro seed during mlanding tempeh’s production.