Comprehensive Comparison of Two Color Varieties of Perillae Folium by GC-MS-Based Metabolomic Approach

Abstract

Perillae Folium (PF), the leaf of Perilla frutescens (L.) Britt, is extensively used as culinary vegetable in many countries. It can be divided into two major varietal forms based on leaf color variation, including purple PF (Perilla frutescens var. arguta) and green PF (P. frutescens var. frutescens). The aroma of purple and green PF is discrepant. To figure out the divergence of chemical composition in purple and green PF, gas chromatography-tandem mass spectrometry (GC-MS) was applied to analyze compounds in purple and green PF. A total of 54 compounds were identified and relatively quantified. Multivariate statistical methods, including principal component analysis (PCA), orthogonal partial least-squares discrimination analysis (OPLS-DA) and clustering analysis (CA), were used to screen the chemical markers for discrimination of purple and green PF. Seven compounds that accumulated discrepantly in green and purple PF were characterized as chemical markers for the discrimination of the purple and green PF. Among these 7 marker compounds, limonene, shisool and perillaldehyde that from the same branch of the terpenoid biosynthetic pathway were with relatively higher contents in purple PF, while perilla ketone, isoegomaketone, tocopheryl and squalene on other branch pathways were higher in green PF. The results of the present study are expected to provide theoretical support for the development and utilization of PF resources.

Keywords: GC-MS; biosynthetic pathway; chemical composition; multivariate statistical analysis; perilla leaf.

Figure 1

The typical total ion chromatograms of n-hexane extracts of (A) purple Perillae Folium and (B) green Perillae Folium by GC-MS. The number of peaks was consistent with those of compounds in Table 2.

Figure 2

Determination of differential compounds from two PF varieties. (A) Unsupervised PCA score plot of purple and green PF samples. PC1 occupies 49.0% and PC2 19.5% of total variance. (B) Supervised OPLS-DA score plot of purple and green PF samples. PC1 occupies 81.5% and PC2 6.73% of total variance. (C) Permutation test at 200 times used for the discrimination between the two PF varieties. (D) Scatter plot of p-value and VIP value. The green points show differential compounds with VIP > 1, p < 0.05.

Figure 3

The relative concentration trends of identified compounds in purple PF and green PF.

Figure 4

Putative biosynthetic pathways of the main terpenoids in perilla. Metabolites are written in black letters, whereas enzymes are written in red letters. DMD, diphosphomevalonate decarboxylase; FDPS, farnesyl diphosphate synthase; LS, limonene synthase; LHS, limonene hydroxylase; PAD, perillylalcohol dehydrogenase; GDD, geranyl diphosphate diphosphohydrolase; FDS, farnesyl diphosphate synthase; SQS, squalene synthase; GGR, geranylgeranyl reductase; TPC, tocopherol C-methyltransferase. *** p < 0.001.