Effects of bilirubin and its photoisomers on direct bilirubin measurement using bilirubin oxidase.
We examined the reactivity of human serum albumin-bound bilirubin and its photoisomers as substrates for a direct bilirubin assay using bilirubin oxidase. The reduction of (EZ)-cyclobilirubin reached 100% 5 min after addition of the enzyme at any pH tested (3.5-7.4) in 0.1 mol/L phosphate buffer, whereas the reduction of (ZE)-bilirubin or (ZZ)-bilirubin reached 100% only below pH 4.5 or 5.5, respectively. (ZZ)-Bilirubin and its photoisomers did not react in citrate-lactate buffer at pH 3.7. The circular dichroism spectrum of (ZZ)-bilirubin in this buffer did not show a positive Cotton effect. These results indicate that a three-dimensional structure surrounding the reaction site of bilirubin is important for the reactivity with bilirubin oxidase.
Itoh
S
S
Department of Pediatrics, Kagawa Medical University, Kitagun, Japan. sonishi@kms.ac.jp
Kusaka
T
T
Imai
T
T
Isobe
K
K
Onishi
S
S
D001663:Bilirubin / Q000097:blood* / Q000737:chemistry / Q000378:metabolism; D002851:Chromatography, High Pressure Liquid; D002942:Circular Dichroism; D019343:Citric Acid / Q000378:metabolism; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007536:Isomerism; D019344:Lactic Acid / Q000378:metabolism; D008027:Light; D008956:Models, Chemical; D010088:Oxidoreductases / Q000737:chemistry / Q000378:metabolism*; D044925:Oxidoreductases Acting on CH-CH Group Donors; D012709:Serum Albumin / Q000378:metabolism*; D013053:Spectrophotometry
D012709:Serum Albumin; D019343:Citric Acid; D019344:Lactic Acid; D010088:Oxidoreductases; D044925:Oxidoreductases Acting on CH-CH Group Donors; C042401:bilirubin oxidase; D001663:Bilirubin