Experiment Center for Teaching and Learning, Shanghai University of Traditional Chinese Medicine, Shanghai 201203, China.
School of Pharmacy, Fudan University, Shanghai 200032, China.
J Zhejiang Univ Sci B. 2019;20(3):264-272. doi: 10.1631/jzus.B1800416.
β-Glucosidase activity assays constitute an important indicator for the early diagnosis of neonatal necrotizing enterocolitis and qualitative changes in medicinal plants. The drawbacks of the existing methods are high consumption of both time and reagents, complexity in operation, and requirement of expensive instruments and highly trained personnel. The present study provides a simplified, highly selective, and miniaturized glucometer-based strategy for the detection of β-glucosidase activity. Single-factor experiments showed that optimum β-glucosidase activity was exhibited at 50 °C and pH 5.0 in a citric acid-sodium citrate buffer when reacting with 0.03 g/mL salicin for 30 min. The procedure for detection was simplified without the need of a chromogenic reaction. Validation of the analytical method demonstrated that the accuracy, precision, repeatability, stability, and durability were good. The linear ranges of β-glucosidase in a buffer solution and rat serum were 0.0873-1.5498 U/mL and 0.4076-2.9019 U/mL, respectively. The proposed method was free from interference from β-dextranase, snailase, β-galactosidase, hemicellulase, and glucuronic acid released by baicalin. This demonstrated that the proposed assay had a higher selectivity than the conventional dinitrosalicylic acid (DNS) assay because of the specificity for salicin and unique recognition of glucose by a personal glucose meter. Miniaturization of the method resulted in a microassay for β-glucosidase activity. The easy-to-operate method was successfully used to detect a series of β-glucosidases extracted from bitter almonds and cultured by Aspergillus niger. In addition, the simplified and miniaturized glucometer-based assay has potential application in the point-of-care testing of β-glucosidase in many fields, including medical diagnostics, food safety, and environmental monitoring.
β-葡萄糖苷酶活性测定是新生儿坏死性小肠结肠炎早期诊断和药用植物定性变化的重要指标。现有方法的缺点是既费时又费试剂，操作复杂，需要昂贵的仪器和经过高度训练的人员。本研究提供了一种简化的、高选择性的、基于血糖仪的策略，用于检测β-葡萄糖苷酶活性。单因素实验表明，在柠檬酸-柠檬酸钠缓冲液中，当与 0.03 g/mL 的水杨苷反应 30 min 时，在 50°C 和 pH 5.0 下，β-葡萄糖苷酶表现出最佳活性。检测过程简化，无需显色反应。分析方法的验证表明，该方法具有良好的准确性、精密度、重复性、稳定性和耐用性。缓冲溶液和大鼠血清中β-葡萄糖苷酶的线性范围分别为 0.0873-1.5498 U/mL 和 0.4076-2.9019 U/mL。该方法不受β-葡聚糖酶、蜗牛酶、β-半乳糖苷酶、木聚糖酶和黄芩苷释放的葡萄糖醛酸的干扰。这表明，由于对水杨苷的特异性和血糖仪对葡萄糖的独特识别，与传统的 3,5-二硝基水杨酸（DNS）法相比，该测定法具有更高的选择性。该方法的微型化导致了β-葡萄糖苷酶活性的微量测定。这种易于操作的方法成功地用于检测从苦杏仁中提取的一系列β-葡萄糖苷酶，并通过黑曲霉进行培养。此外，简化和微型化的血糖仪测定法在许多领域，包括医学诊断、食品安全和环境监测，都具有在β-葡萄糖苷酶的即时检测中的潜在应用。
