The enantioselective immunoaffinity extraction of an optically active ibuprofen-modified peptide fragment
The enantioselective immunoaffinity extraction of an optically active ibuprofen-modified peptide fragment
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DOI：
10.1006/abio.2001.5198
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发表时间：
2001-09-01
影响因子：
2.9
通讯作者：
Goto, J
中科院分区：
文献类型：
--
作者：
Ikegawa, S;Isriyanthi, NMR;Goto, J
已知酰基葡萄糖醛酸苷可产生共价结合的蛋白加合物，这可能是对酸性药物产生超敏反应和毒性反应的原因。因此，需要对药物-蛋白质加合物进行结构分析。从这一点来看，我们开发了一种对映选择性免疫亲和提取方法，该方法采用固定化抗体来特异性地分离已经用光学活性布洛芬修饰的肽片段。用通过β-丙氨酸基团偶联到牛血清白蛋白的（S）-布洛芬免疫家兔。所引发的抗体强烈识别（S）-布洛芬及其缀合物的不对称中心和异丁基苯基部分，但对其反足具有低亲和力。通过固定抗体制备的0.5 mL免疫吸附剂等分试样（11.5 mg IgG/mL凝胶）能够保留高达1 μ g的（S）-布洛芬。当物质P与（R）-和（S）-布洛芬修饰的物质P的混合物被加载到免疫吸附剂上时，（S）-布洛芬修饰的物质P被选择性地保留。通过用10 mM乙酸铵缓冲液（pH 5.0）/甲醇（5：95，v/v）洗脱定量回收修饰的肽。该方法可用于光学活性布洛芬修饰人血清白蛋白的结构表征。(C)北京：科学出版社.
Acyl glucuronides are known to produce the covalently bound protein adducts which may be the cause of hypersensitivity and toxic responses to acidic drugs. The structural analysis of the drug-protein adducts is therefore needed. From this point of view, we developed an enantioselective immunoaffinity extraction method, which employs an immobilized antibody to specifically isolate peptide fragments that have been modified with optically active ibuprofen. Rabbits were immunized with (S)-ibuprofen coupled to bovine serum albumin through a beta -alanine group. The elicited antibody strongly recognizes the asymmetric center and the isobutylphenyl moiety of (S)-ibuprofen and its conjugates but has a low affinity for their anti podes. A 0.5-mL aliquot of the immunosorbent (11.5 mg of IgG/mL gel) prepared by immobilization of the antibody was capable of retaining up to 1 mug of (S)-ibuprofen. When a mixture of substance P with (R)- and (S)-ibuprofen-modified substance P was loaded on the immunosorbent, the (S)-ibuprofen-modified substance P was selectively retained. The modified peptide was quantitatively recovered by elution with 10 mM ammonium acetate buffer (pH 5.0)/methanol (5:95, v/v). The proposed method would be useful for the structural characterization of optically active ibuprofen-modified human serum albumin. (C) 2001 Academic Press.