Blasi Francesca, Ianni Federica, Mangiapelo Luciano, Pinna Nicola, Cossignani Lina
Department of Pharmaceutical Sciences, University of Perugia, Perugia, Italy.
Center for Perinatal and Reproductive Medicine, Santa Maria della Misericordia University Hospital, University of Perugia, Perugia, Italy.
J Sci Food Agric. 2023 Apr;103(6):2786-2793. doi: 10.1002/jsfa.12417. Epub 2023 Jan 12.
Pancreatic lipase (PL) is a key lipolytic enzyme in humans for the digestion and absorption of dietary fats. Thereby, PL is a well-recognized target in the management of obesity and its inhibition attracts the interest of researchers globally. The screening of new natural PL inhibitors as alternative strategy to the synthesis of chemical ones represents nowadays a hot topic in research. The main challenge in this matter is the lack of a universal analytical method allowing the monitoring of PL activity and the reliable quantification of lipid digestion products.
The (normal phase)-high-performance liquid chromatography-evaporative light scattering detector [(NP)-HPLC-ELSD] method proposed in this work represents a direct and rapid strategy to simultaneously quantify the products obtained from in vitro PL digestion. As one of the main novelties, the triacylglycerol (TAG) fraction from extra-virgin olive oil was selected as natural substrate. The PL activity was measured by monitoring the levels of remaining TAGs and formed free fatty acids (FFAs), using Orlistat as known inhibitor. The method validation confirmed the adequacy of the analytical method for quantitative purposes, showing high recovery percentage values (between 99% and 103%) and low relative standard deviation (RSD%) values (between 2% and 7%) for triolein and oleic acid standard solutions, as well as appreciably low limit of detection (LOD) and limit of quantification (LOQ) values (respectively 58 and 177 ng mL for triolein; 198 and 602 ng mL for oleic acid). Finally, the developed HPLC-ELSD method was successfully applied to evaluate the inhibitory effect of a polyphenolic extract obtained from apple pomace. The results showed a comparable inhibition degree between a 4.0 mg mL apple pomace solution and a 1.0 μg mL Orlistat solution.
The proposed innovative method reveals highly sensitive and simple to follow the fate of PL digestion, thus opening the way to further investigations in the research of new potentially anti-obesity compounds. © 2022 Society of Chemical Industry.
胰腺脂肪酶（PL）是人体中负责消化和吸收膳食脂肪的关键脂解酶。因此，PL是肥胖管理中一个公认的靶点，其抑制作用吸引了全球研究人员的关注。筛选新型天然PL抑制剂作为合成化学抑制剂的替代策略，是当前研究的热点话题。这方面的主要挑战是缺乏一种通用的分析方法来监测PL活性并可靠地定量脂质消化产物。
本研究提出的（正相）高效液相色谱 - 蒸发光散射检测器[（NP）-HPLC-ELSD]方法是一种直接且快速的策略，可同时定量体外PL消化获得的产物。作为主要创新之一，特级初榨橄榄油中的三酰甘油（TAG）部分被选作天然底物。以奥利司他作为已知抑制剂，通过监测剩余TAG和生成的游离脂肪酸（FFA）水平来测定PL活性。方法验证证实了该分析方法用于定量目的的适用性，三油酸甘油酯和油酸标准溶液的回收率较高（99%至103%），相对标准偏差（RSD%）较低（2%至7%），检测限（LOD）和定量限（LOQ）值也相当低（三油酸甘油酯分别为58和177 ng/mL；油酸为198和602 ng/mL）。最后，所开发的HPLC-ELSD方法成功应用于评估苹果渣多酚提取物的抑制作用。结果表明，4.0 mg/mL的苹果渣溶液与1.0 μg/mL的奥利司他溶液的抑制程度相当。
所提出的创新方法在跟踪PL消化过程方面显示出高度灵敏且简单，从而为进一步研究新型潜在抗肥胖化合物开辟了道路。© 2022化学工业协会。
