## 3 Results and Discussion  
### UPLC-QTOF/MS with an In-House Library to Profile Various Ginsenosides  
The sensitivity of MS-based analytical methods has increased to enable the detection of compounds with limited abundance in biological samples. Thus, LC-MS methods have been widely used to analyze major compounds in various herbal medicines. For the detailed characterization of _P. ginseng_, assessing the levels of both the high- and low-abundance ginsenosides is important. In our previous study [25], UPLC-QTOF/MS was used to analyze 30 ginsenosides. The optimal UPLC method was effective for separating various ginsenosides. In the identification of ginsenosides, QTOF/MS was also sensitive and reliable for measuring the exact mass values of the compounds. In this study, the LC-MS method was simultaneously applied to profile 58 ginsenosides. Figure S1 represented the molecular structures of these ginsenosides. First, these ginsenoside standards were purchased or obtained from the experiments by isolating and purifying them from _P. ginseng_. In our laboratory, we collected four MGRs that contained a malonyl residue, M-Rb1, M-Rb2, M-Rc, and M-Rd, which is linked to the glucose in the compounds. Isolating and purifying the MGRs from the extracted mixture of _P. ginseng_ is difficult in practice. Due to their instability when exposed to heat, care must be taken when handling and preserving MGR compounds.  
Next, for the comprehensive profiling of various ginsenosides, 58 ginsenoside standards were used to optimize the UPLC-QTOF/MS conditions. By using the UPLC system with an ACQUITYBEH C18 column, these standards were well separated after 30 min at a flow rate of 450 \(\mu\)L/min. The base peak intensity (BPI) chromatograms of the 58 ginsengides are shown in Figure 1. Among these ginsengides, 16 isomeric well-separated pairs were found: C\({}_{36}\)H\({}_{60}\)O\({}_{7}\) (Rk2 or Rh3), C\({}_{36}\)H\({}_{60}\)O\({}_{8}\) (Rk3, Rh4, fKa, or Rh6), C\({}_{36}\)H\({}_{62}\)O\({}_{8}\) (Rh2 or CK), C\({}_{36}\)H\({}_{62}\)O\({}_{9}\) (Rh1, Rh1-R, or F1), C\({}_{41}\)H\({}_{70}\)O\({}_{12}\) (Mc or CY), C\({}_{41}\)H\({}_{70}\)O\({}_{13}\) (nR2, F5, nR2-R, or F3), C\({}_{42}\)H\({}_{70}\)O\({}_{12}\) (Rg4, F4, Rk1, or Rg5), C\({}_{42}\)H\({}_{72}\)O\({}_{13}\) (Rg2, Rg2-R, F2, Rg3, or Rg3-R), C\({}_{42}\)H\({}_{72}\)O\({}_{14}\) (Rg1, pF11, Rf, GyL, or GyLLI), C\({}_{47}\)H\({}_{80}\)O\({}_{17}\) (nFe, CO, or nFt1), C\({}_{48}\)H\({}_{82}\)O\({}_{18}\) (Re, Rd, or GyXVII), C\({}_{48}\)H\({}_{82}\)O\({}_{19}\) (gRf or vR4), C\({}_{53}\)H\({}_{90}\)O\({}_{22}\) (Rc, Rb2, or Rb3), C\({}_{56}\)H\({}_{92}\)O\({}_{25}\) (M-Rc or M-Rb2), C\({}_{58}\)H\({}_{98}\)O\({}_{26}\) (Ra1 or Ra2), and C\({}_{59}\)H\({}_{100}\)O\({}_{27}\) (nR4 or Ra3). Although several peaks overlapped with each other, identifying each peak was possible, based on its exact mass values obtained by QTOF/MS. In the negative ion mode, ginsengides were detected as [M + COOH]\({}^{-}\) ions and [M - H]\({}^{-}\) ions with high mass accuracy (<2.5 ppm). For the exact identification of ginsengides, we constructed an in-house library using the UNIFI software. Information about the compound's name, molecular formula, retention time (RT), mass accuracy, and adduct was listed in the library for the 58 ginsengides (Table 1). Next, a validation study was performed to prove the performance of the UPLC-QTOF/MS method for profiling the ginsengides. For this, the standard curves of the 58 ginsengides were constructed, and their linearity range and correlation (\(R^{2}\)) were calculated. The limits of detection (LODs) of each standard are provided in Table S1. The \(R^{2}\) of each ginsengide analysis was, at minimum, above 0.9908, proving the high reliability of the quantitative analysis of the ginsengides. LODs also showed the high sensitivity of this method for detecting ginsengides. For example, in the case of Ra3, Ro, M-Rb2, Rd, GyL, GyLLI, Rg3, Mc, and Cy, the two picogram (pg) of these compounds was detected by UPLC-QTOF/MS.