In the present study, ammonium sulfate fractional precipitation, PEG 6000 precipitation and Sephadex G-200 column chromatography were sequentially carried out to separate and purify alliinase from crude garlic extract. SDS-PAGE was used to identify the enzyme and analyze its purity. The optimal extraction and separation conditions were determined as follows： alliinase was extracted with pH 6.5 phosphate buffer at 4 ℃, precipitated with 20% PEG 6000, and centrifugated at 2000 μg for 30 min. After purification by Sephadex G-200 column chromatography, the purification factor of the purified alliinase relative to the crude alliinase extract was 16.85 fold and activity recovery 41.26%. The subunit molecular weight of the alliinase was approximately 54.7 kD. An orthogonal array design was use to optimize the effects of three metal ions, glucose and sucrose on its activity. Moreover, the effect of metal ions on its reaction kinetics and UV-visible spectrum was also explored. The results showed that combinations of metal ions at various concentrations revealed a significant impact on alliinase activity. Mn2. was the most important factor. In the presence of 10 mmol/L Mn^2＋, 10 mmol/L Fe^3＋, 5 mmol/L Ca^2＋, and 5 mmol/L sucrose, the highest relative activity of 155.59% was obtained. However, 10 mmol/L Cu^2＋ significantly inhibited alliinase activity. The Km was measured to be 0.25 mmol/L and Vmax was determined to be 1.29 μmol/min using alliin as the substrate. UV-visible spectroscopy was used to confirm the existence of the cofactor pyridoxal phosphate （PLP）. The activation or inhibition of aUiinase by metal ions may be due to changes in the structure of 5＇-PLP and its binding to the main chain of alliinase.%以新鲜大蒜为原料，采用硫酸铵分级沉淀法、PEG6000沉淀法、葡聚糖G-200柱层析法提取分离蒜氨酸酶，获得适宜蒜氨酸酶粗分离的条件为：4℃、pH6．5磷酸盐缓冲液浸提，20％的PGE6000盐析，2000×g离心30min。再采用SDS—PAGE电泳法鉴定其纯度，发现纯度是粗酶液的16．85倍，回收率为41．26％，蒜氨酸酶亚基的分子质量约为54．7kD。利用正交试验优化金属离子交互作用对蒜氨酸酶活性的影响，研究金属离子对蒜氨酸酶反应动力学及紫外-可见光谱的影响。结果表明：不同浓度的离子组合对蒜氨酸酶活性有显著的影响，Mn^2＋是影响蒜氨酸酶活性最重要的因素；不同离子交互作用最佳条件为：Mn^2＋浓度10mmol／L、Fe^3＋浓度10mmol／L、Ca^2＋浓度5mmol／L、辅助因子蔗糖浓度5mmol／L，此条件下蒜氨酸酶相对酶活性为155．59％。而10mmol／L的Cu^2＋能够显著抑制蒜氨酸酶的活性；以蒜氨酸为底物，测得蒜氨酸酶蠡Km值为0．25mmol／L，％。为1．29μmol／min；紫外-可见光谱证实了辅基磷酸吡哆醛的存在，金属离子对蒜氨酸酶的激活与抑制机制可能是因为激活剂和抑制剂影响了蒜氨酸酶辅助因子5’-磷酸吡哆醛的结构以及其与蒜氨酸酶主链的结合，从而影响了蒜氨酸酶的活性。
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