Zhou Guisheng, Wang Mengyue, Li Yang, Peng Ying, Li Xiaobo
School of Pharmacy, Shanghai Jiao Tong University, Shanghai, 200240, China.
Amino Acids. 2015 Aug;47(8):1589-603. doi: 10.1007/s00726-015-2002-5. Epub 2015 May 15.
In the present study, a new strategy based on chemical analysis and chemometrics methods was proposed for the comprehensive analysis and profiling of underivatized free amino acids (FAAs) and small peptides among various Luo-Han-Guo (LHG) samples. Firstly, the ultrasound-assisted extraction (UAE) parameters were optimized using Plackett-Burman (PB) screening and Box-Behnken designs (BBD), and the following optimal UAE conditions were obtained: ultrasound power of 280 W, extraction time of 43 min, and the solid-liquid ratio of 302 mL/g. Secondly, a rapid and sensitive analytical method was developed for simultaneous quantification of 24 FAAs and 3 active small peptides in LHG at trace levels using hydrophilic interaction ultra-performance liquid chromatography coupled with triple-quadrupole linear ion-trap tandem mass spectrometry (HILIC-UHPLC-QTRAP(®)/MS(2)). The analytical method was validated by matrix effects, linearity, LODs, LOQs, precision, repeatability, stability, and recovery. Thirdly, the proposed optimal UAE conditions and analytical methods were applied to measurement of LHG samples. It was shown that LHG was rich in essential amino acids, which were beneficial nutrient substances for human health. Finally, based on the contents of the 27 analytes, the chemometrics methods of unsupervised principal component analysis (PCA) and supervised counter propagation artificial neural network (CP-ANN) were applied to differentiate and classify the 40 batches of LHG samples from different cultivated forms, regions, and varieties. As a result, these samples were mainly clustered into three clusters, which illustrated the cultivating disparity among the samples. In summary, the presented strategy had potential for the investigation of edible plants and agricultural products containing FAAs and small peptides.
在本研究中，提出了一种基于化学分析和化学计量学方法的新策略，用于对不同罗汉果（LHG）样品中的未衍生化游离氨基酸（FAA）和小肽进行综合分析和表征。首先，使用Plackett-Burman（PB）筛选和Box-Behnken设计（BBD）对超声辅助提取（UAE）参数进行优化，得到以下最佳UAE条件：超声功率280W、提取时间43分钟、固液比302mL/g。其次，开发了一种快速灵敏的分析方法，使用亲水相互作用超高效液相色谱-三重四极杆线性离子阱串联质谱（HILIC-UHPLC-QTRAP(®)/MS(2)）同时定量LHG中痕量水平的24种FAA和3种活性小肽。通过基质效应、线性、检测限、定量限、精密度、重复性、稳定性和回收率对该分析方法进行了验证。第三，将所提出的最佳UAE条件和分析方法应用于LHG样品的测定。结果表明，LHG富含必需氨基酸，这些是对人体健康有益的营养物质。最后，基于27种分析物的含量，采用无监督主成分分析（PCA）和有监督的反向传播人工神经网络（CP-ANN）化学计量学方法对40批来自不同栽培形式、地区和品种的LHG样品进行区分和分类。结果，这些样品主要聚为三类，这说明了样品之间的栽培差异。总之，所提出的策略在含FAA和小肽的可食用植物和农产品研究方面具有潜力。
