凝集素 (lectin) 是一種具有醣類結合專一性之蛋白或醣蛋白，且因具有醣類專一性，所以能使紅血球發生凝集現象，也同時具有許多生理功能。本研究目的是將胡蘿蔔食用肉根部分的植物凝集素根據carbohydrate-binding specificity of the carrot lectin中所採用之萃取條件，利用管柱層析法將胡蘿蔔凝集素分離純化。另外也用其他兩種不同條件pH值之buffer進行萃取，比較其凝集活性。 純化部分，由實驗結果得知，可在硫酸銨飽和度達45% - 95%區間內得到較高比活性之蛋白質。經由陰離子交換層析管柱進一步純化，當鹽度達65-100%時可沖提出之蛋白質具有凝集活性。再利用親和性管柱進行下一步純化。 而萃取條件比較方面，利用5%醋酸（pH 2.5）、PBS buffer (pH 7.4) 和carbonate - bicarbonate buffer (pH 10.5) 三種條件，對 40 g 胡蘿蔔食用的根進行萃取。由實驗結果顯示，以carbonate - bicarbonate buffer 萃取之萃取物可得到最大之比活性。隨後比較N-acetyl-D-glucosamine、D(+)-glucose、D(+)-mannose、D(+)-galactose、maltose、D(+)-glucosamine、D(-)-fructose、saccharose 八種醣類對萃取物之活性抑制情形。結果顯示，雖然這八種醣類皆無法對各種萃取物之活性產生特異性之抑制，或許能夠提供將來純化之胡蘿蔔凝集素進一步研究之方向。 Lectins are proteins which can agglutinate erythrocytes, not only can bind carbohydrates, also have some specific biological activities. In this study we extracted carrot root lectin and used precipitation method and chromatography to purify carrot root lectin. The extraction was also carried out in different pH buffers, looking for the effect of pH on hemagglutinating activities. For purification, we extracted the lection activity by following Carbohydrate-binding Specificity of the Carrot .Ammonium sulfate fraction at 45 - 95% recovered the highest specific activity. For ion exchanger column, buffer B at 65 - 100 %, could elute the agglutinin activity. Finally, the affinity column could further purify carrot root lectin. For the extraction step, we chose two other pH buffers: PBS buffer (pH 7.4) and Carbonate - Bicarbonate buffer (pH 10.5), and compared the effect with 5% acetate (pH 2.5), hoping to find out distinct hemagglutinating activities and carbohydrate’s specificities.
