Sonesson B, Eide S, Rorsman H, Rosengren E
Department of Dermatology, University of Lund, Sweden.
Acta Derm Venereol. 1995 Jul;75(4):283-6. doi: 10.2340/0001555575283286.
Serum samples for the analysis of tyrosinase activity were obtained from 10 healthy subjects in autumn, winter and summer. Tyrosinase was purified from 100 microl serum by adsorption to concanavalin A sepharose, the tyrosinase adsorbed to the gel being separated from other components by centrifugation. The gel was suspended in a buffer containing 5-hydroxy-indole-3-acetic acid as an antioxidant and incubated for 2 min with L-cysteine and D-L-dopa at 37 degrees C. The 5-S-L-cysteinyl-L-dopa formed was measured by HPLC and electrochemical detection. Tyrosinase has high stereo-specificity for the L-enantiomer of dopa, and correction for non-specific oxidation was made by simultaneous measurement of 5-S-L-cysteinyl-D-dopa formed from D-dopa. Whereas the oxidation of L-dopa catalysed by tyrosinase ws inhibited by L-tyrosine, the non-specific oxidation of D-dopa was not. Mean serum tyrosinase activity was 0.9 nkatal/l in summer, 0.8 nkatal/l in autumn and 0.4 nkatal/l in winter. The range of tyrosinase activity was much higher in summer and autumn than in winter.
为分析酪氨酸酶活性，分别在秋季、冬季和夏季从10名健康受试者身上采集血清样本。通过吸附到伴刀豆球蛋白A琼脂糖上从100微升血清中纯化酪氨酸酶，通过离心将吸附到凝胶上的酪氨酸酶与其他成分分离。将凝胶悬浮在含有5-羟基吲哚-3-乙酸作为抗氧化剂的缓冲液中，并在37℃下与L-半胱氨酸和D-L-多巴孵育2分钟。通过高效液相色谱法和电化学检测法测定形成的5-S-L-半胱氨酰-L-多巴。酪氨酸酶对多巴的L-对映体具有高度立体特异性，通过同时测量由D-多巴形成的5-S-L-半胱氨酰-D-多巴来校正非特异性氧化。虽然酪氨酸酶催化的L-多巴氧化受到L-酪氨酸的抑制，但D-多巴的非特异性氧化不受抑制。夏季血清酪氨酸酶平均活性为0.9纳 katal/升，秋季为0.8纳 katal/升，冬季为0.4纳 katal/升。夏季和秋季酪氨酸酶活性范围比冬季高得多。
