环丙沙星衍生物的制备及其抗体特异性研究

本文研究了环丙沙星(CPLX)羧基衍生物及其抗体的制备,并对该衍生物抗体的特异性进行了测定。将环丙沙星与3-溴丙酸反应,生成环丙沙星的羧基衍生物(CPLX-COOH),通过制备液相进行纯化,用红外光谱(IR)、电喷雾电离质谱(ESI-MS)与核磁共振法(NMR)进行鉴定;将环丙沙星衍生物分别与牛血清白蛋白(BSA)和卵清蛋白(OVA)进行偶联,分别制备免疫抗原(CPLX-COOH-BSA)和包被抗原(CPLX-COOH-OVA),并用紫外光谱法进行鉴定。用免疫抗原免疫BALB/c小鼠,制备环丙沙星衍生物多克隆抗体。通过ELISA方阵滴定法,确定包被抗原的最佳包被浓度和抗体最佳稀释度,建立了CPLX-COOH间接竞争ELISA检测方法,其半抑制浓度IC50为228.56 ng/mL,最低检出限(LOD)为25.527 ng/mL,该抗体具有广谱特异性,可与多种氟喹诺酮类药物发生交叉反应,可进行氟喹诺酮类药物残留的多元检测。

Preparation of Ciprofloxacin Derivative and its Antibody Specificity

The preparations of the ciprofloxacin (CPLX) carboxyl derivative and its antibody were studied in this paper, and the antibody specificity of this derivative was determined. CPLX derivatives (CPLX-COOH) were synthesized with CPLX and 3-bromopropionic acid, and purified by preparation of liquid phase. Infrared spectroscopy (IR), electrospray ionization mass spectrometry (ESI-MS), and nuclear magnetic resonance (NMR) spectroscopy were used to identify the CPLX derivative. CPLX-COOH was conjugated to bovine serum albumin (BSA) and ovalbumin (OVA) to prepare the immune antigen CPLX-COOH-BSA and coating antigen CPLX-COOH-OVA, which were identified using ultraviolet spectroscopy. The BALB/c mice were immunized with CPLX-COOH-BSA to obtain a polyclonal antibody directed against the CPLX derivative with high titer. The optimum concentration of the coating antigen and the optimum dilution of the antibody were determined using the chessboard method, and the indirect competitive enzyme-linked immunosorbent assay (ELISA) was established. The half-maximal inhibitory concentration (IC50) and the limit of detection (LOD) of CPLX-COOH were 228.56 ng/mL and 25.527 ng/mL, respectively. This antibody had a broad specificity and cross reactivity with multiple fluoroquinolones, and could be used to detect several fluoroquinolone residues.