원유중 잔류 설파제의 검출을 위한 2,3,5-triphenyltetrazolium chloride(TTC) 환원시험법의 개량

Development of modified 2, 3, 5-triphenyltetrazolium choride(TTC) reduction test for the detection of sulfonamide residues in raw milk

A modified 2,3,5-triphenyltetrazolium chloride(TTC) reduction test for the detection of sulfonamide residues in raw milk is presented. The test is based on the conversion of TTC to formazan mediated by the enzyme preduced from Streptococcus thermophilus ATCC 14485. The reaction is characterized by a color change from white to red in the presence of growing test organism. When a sulfonamide is combined with trimethoprim(TMP), a synergistic effect on microbial inhibition is exerted. On the other hand, p-aminobenzoic acid (PABA) blocks the inhibitory effect of sulfonamides on bacterial growth. For this reason, 5.0 ㎍ of TMP and 50 ㎍ of PABA per milliliter were added to milk samples to improve the detection limits of sulfonamide residues and also to differentiate sulfonamide from other antimicrobial drug residues in the modified test. When the detection limits of sulfonamide residues in raw milk were tested by the modified method, the levels of sulfachloropyridazine, sulfadimethoxine, sulfadiazine, sulfamonomethoxine, sulfaquinoxaline, sulfisomidine, and sulfathiazole at 0.025 ppm and of sulfamerazine, sulfamethazine, and sulfamethoxypyridazine at 0.05 ppm could be detected, respectively. The results suggested that the minimum detectable levels of sulfonamide residues were significantly improved. In addition, seven β-lactams at 0.005-0.05 ppm, three tetracyclines at 0.1-0.25 ppm, four macrolides at 0.25 ppm, three aminoglycosides at 5.0-10.0 ppm, chloramphenicol at 0.1 ppm, and bacitracin at 0.25 ppm could also be detected in raw milk by the modified test. Therefore, the modified TTC reduction test could be applied as a screening method for the detection of antimicrobal drug residues in raw milk.