The promotion of hexadecane biodegradation activity by an n-alkane degrading strain of Burkholderia cepacia (GS3C) with yeast extract amendment was studied using various carbon, nitrogen, vitamin, and amino acid amendments. Cytochrome P450 monooxygenase enzymes play a very important role and are especially required to introduce oxygen in n-alkane degradation. These enzymes from GS3C were located and detected using amino acid amendments. It was shown that biodegradation activity was promoted with amino acids amendments. However, only specific amino acids (l-phenylalanine, l-glutamic acid, l-proline, l-lysine, l-valine and l-leucine) have biodegradation promoting ability for GS3C. Cell protein concentration and cytochrome P450 activity were promoted significantly with the addition of l-phenylalanine and yeast extract. Furthermore, a significant positive linear relationship between cytochrome P450 activity and biodegradation efficiency of GS3C was observed. The results indicate that amino acid is the primary factor of nutrient amendment in promoting hexadecane biodegradation by influencing cytochrome P450 activity in GS3C.