School of Food Science, Washington State University , Pullman, Washington 99164-6376, United States.
J Nat Prod. 2013 Sep 27;76(9):1605-11. doi: 10.1021/np400264b. Epub 2013 Aug 21.
Vaccinium macrocarpon (cranberry) products have been used to prevent uropathogenic Escherichia (E.) coli adherence to uroepithelial cells (UEC) and may help reduce risk of urinary tract infection. Reported herein are the development and validation of an assay to assess antiadhesion activity of V. macrocarpon extracts and human urine. P-fimbriated E. coli (CFT073) was labeled with ³H-uridine, then co-incubated with HTB-4 UEC at a 400:1 ratio. V. macrocarpon extracts (0-17 mg proanthocyanidins/mL) were added to ³H-labeled E. coli before co-incubating with UEC. The assay yielded a sensitive inhibition curve: the lower limit of detection and half-maximal inhibitory concentration were 0.43 and 1.59 mg proanthocyanidins/mL for V. macrocarpon extract CEP 55; intra- and interassay coefficients of variance were <10% and <15%, respectively. V. macrocarpon extract CEP 3283 showed identical adhesion inhibition. Serial dilutions of urine from human participants who consumed V. macrocarpon beverages showed a linear decrease in antiadhesion activity. Antiadhesion assays conducted with urine from a human intervention study also showed good agreement with results obtained using the hemagglutination assay. Therefore, a sensitive, high-throughput, biologically relevant antiadhesion assay using ³H-E. coli co-incubated with UEC is reported, which can be used for studying the action of V. macrocarpon bioactives.
蔓越莓（Vaccinium macrocarpon）产品已被用于预防尿路致病性大肠杆菌（Escherichia coli）黏附于尿路上皮细胞（uroepithelial cells，UEC），并可能有助于降低尿路感染的风险。本文报告了一种评估蔓越莓提取物和人尿液抗黏附活性的分析方法的开发和验证。用 ³H-尿嘧啶对 P-菌毛大肠杆菌（CFT073）进行标记，然后以 400:1 的比例与 HTB-4 UEC 共孵育。将 ³H 标记的大肠杆菌与 UEC 共孵育前，加入蔓越莓提取物（0-17 mg 原花青素/mL）。该分析方法得到了灵敏的抑制曲线：蔓越莓提取物 CEP 55 的检测下限和半抑制浓度分别为 0.43 和 1.59 mg 原花青素/mL；内和间试验变异系数均<10%和<15%。蔓越莓提取物 CEP 3283 显示出相同的黏附抑制作用。来自食用蔓越莓饮料的人类参与者尿液的连续稀释显示出抗黏附活性呈线性下降。使用来自人类干预研究的尿液进行的抗黏附分析也与使用血凝分析获得的结果具有良好的一致性。因此，本文报道了一种使用 ³H-大肠杆菌与 UEC 共孵育的灵敏、高通量、具有生物学相关性的抗黏附分析方法，可用于研究蔓越莓生物活性物质的作用。
