Abstract: Objective To study the effect of D-Tryptophan, D-Leucine, D-tyrosine, and D-Methionine on the formation of biofilm Porphyromonas gingivalis (P. gingivalis) W83. Method: P. gingivalis W83 were cultured in Trypticase soy broth (TSB) supplemented with vitamin K (1μg/ml) and hemin (5 μg/ml) to mid-log phase, and then were diluted 1:100 in 96 well flat-bottom poltstyrene plates and add D-Tryptophan (5mmol/L), D-Leucine (8.5 mmol/L), D-tyrosine (3μmol/L), and D-Methionine (2mmol/L). The biofilms were stained with 25μl of 1% crystal violet, washed with ddH2O and dissolved in ethanol. The absorbance was measured with xMark microplate spectrophotometer at 570nm. Result: The absorbance of P. gingivalis W83 in TSB, D-Tryptophan, D-Leucine, D-tyrosine, and D-Methionine were 3.70 ± 0.06, 2.74 ± 0.18, 0.71 ± 0.04, 3.42 ± 0.02, and 3.65 ± 0.01 respectively. There was significance difference between TSB and D-Leucine groups (p<0.01). Conclusion D-Leucine (8.5 mmol/L) can reduce the biofilm formation of P. gingivalis W83.