Agatonovic-Kustrin Snezana, Morton David W
School of Pharmacy, Monash University Malaysia, Jalan Lagoon Selatan, Bandar Sunway, 47500, Selangor Darul Ehsan, Malaysia.
School of Pharmacy and Applied Science, La Trobe Institute of Molecular Sciences, La Trobe University, Edwards Rd, Bendigo, 3550, Australia.
MethodsX. 2018 Jul 20;5:797-802. doi: 10.1016/j.mex.2018.07.013. eCollection 2018.
A high-performance thin-layer chromatography (HPTLC) method was developed for quantification of α-amylase inhibitory activity and stigmasterol content in ant plant extracts. An improved HPTLC method for the determination of total free radical scavenging activity in samples using DPPH• is also reported. For quantification of -amylase inhibitory activity, the developed HPTLC plate is dipped into an -amylase solution, and the bioautogram is then incubated at 25 °C for 30 min under humid conditions. For visualization of enzyme inhibitory activity, the starch test with an iodine indicator solution is used. The blue zone observed comes from the starch-iodine complex formed from starch that was not hydrolyzed by the amylase due to enzyme inhibition by the compound(s) present in the sample. The area of the blue zones was used to compare and quantify relative α-amylase inhibitory activity in different extracts. Location of the blue zones (hRF) on the plate was used to detect compounds that are responsible for the α-amylase inhibitory activity. Relative α-amylase activity was not related to the antioxidant activity, but was highly correlated with the stigmasterol content in the sample extracts ( = 0.95). Therefore, plant sterols present in the extracts might be responsible for α-amylase inhibitory activities in the extracts. •The developed method for quantification of α-amylase inhibitory activity provides an efficient and effective tool that can be used to screen, detect and quantify α-amylase inhibitory activity in plant extracts.•The proposed protocol is easy to run, involves minimal sample preparation, with multiple samples able to be analyzed in parallel on the same chromatographic plate, in a short time.•There were significant differences in -amylase inhibitory activity, stigmasterol content, and total free radical scavenging activity between methanol, ethanol, dichloromethane, and ethyl acetate ant plant extracts.
开发了一种高效薄层色谱（HPTLC）方法，用于定量蚂蚁植物提取物中的α-淀粉酶抑制活性和豆甾醇含量。还报道了一种改进的HPTLC方法，用于使用DPPH•测定样品中的总自由基清除活性。为了定量α-淀粉酶抑制活性，将开发的HPTLC板浸入α-淀粉酶溶液中，然后在潮湿条件下于25°C孵育生物自显影图谱30分钟。为了可视化酶抑制活性，使用碘指示剂溶液进行淀粉试验。观察到的蓝色区域来自淀粉-碘复合物，该复合物由未被淀粉酶水解的淀粉形成，这是由于样品中存在的化合物对酶的抑制作用。蓝色区域的面积用于比较和定量不同提取物中相对α-淀粉酶抑制活性。板上蓝色区域的位置（比移值）用于检测负责α-淀粉酶抑制活性的化合物。相对α-淀粉酶活性与抗氧化活性无关，但与样品提取物中的豆甾醇含量高度相关（r = 0.95）。因此，提取物中存在的植物甾醇可能是提取物中α-淀粉酶抑制活性的原因。•开发的α-淀粉酶抑制活性定量方法提供了一种高效且有效的工具，可用于筛选、检测和定量植物提取物中的α-淀粉酶抑制活性。•所提出的方案易于操作，样品制备最少，多个样品能够在同一色谱板上并行分析，且耗时短。•甲醇、乙醇、二氯甲烷和乙酸乙酯蚂蚁植物提取物之间的α-淀粉酶抑制活性、豆甾醇含量和总自由基清除活性存在显著差异。
