家蝇源抗K562活性物质的分离与纯化

ISOLATION AND PURIFICATION OF COMPONENTS WITH ANTI-K562 ACTIVITY FROM THE MUSCA DOMESTICA

为从家蝇Ⅲ龄幼虫体内提取具有抗K562细胞活性的物质，并对其组分化学性质及对人正常细胞的作用进行分析。本文用大肠杆菌诱导家蝇Ⅲ龄幼虫大量表达活性物质，然后通过研磨、离心、固相萃取法（SPE）初步分离活性物质，通过反相高效液相色谱（RP—HPLC）进一步纯化提取活性组分，通过四甲基偶氮噻唑蓝（Methyl thiazolyl tetrazolium，MTT）比色法和光镜观察法相结合，筛选出对K562有抑制作用的活性组分，并鉴定其对人正常细胞293T的作用。结果显示，从家蝇Ⅲ龄幼虫体内分离出4个具有抑制K562细胞活性的多肽组分（峰5～8），具有分离度高、纯度高等特点。当质量浓度为100ug／mL和作用时间为24h时，4个多肽组分均具有抗K562细胞的活性，其中峰5多肽组分和峰8多肽组分活性较强，对K562细胞的相对抑制率分别为48．52％和65．80％。结果表明，家蝇Ⅲ龄幼虫体内存在一些具有抗K562细胞活性的多肽，而且对人正常细胞几乎无杀伤作用。但是，这些多肽是否属于抗肿瘤肽还有待被证实。

The purpose of this paper is to extracte biological components with anti-K562 activity from the Musca domestica larvae, and to determine their chemical properties and roles to the normal-human cells preliminarily. Firstly, the activity components from the M. domestica larvae were induced by E. coli and isolated by using trituration, centrifugalization, solid phase extraction （SPE） followed by reversed-phase high-performance liquid chromatography （RP-HPLC）. The anti-cell （to both K562 and normal-human cells of 293T） activity of these components was deteremined by the MTT colorimetric method and light microscope. Four polypeptide components （peaks 5 to 8 ） with anti-KS62 activities were isolated from the M. domestica Larvae. When the concentrations was 100 ~g/mL and the reaction time was 24 h, the four components showed anti-K562 activity and had almost no damage effect to normal-human cells. Of these components, peak 5 and peak 8 had stronger inhibition activity, with IRs （the relative inhibition rate） of 48.52% and 65.80% respectively.