Application of the new at-column dilution (ACD) modulator for the two-dimensional RP×HILIC analysis of Buddleja davidii

Abstract

The focus of this study was the analysis of the complex chemical composition from different parts of Buddleja davidii, whose species are commonly known as ornamental plants and herbal medicines in many countries. As an herbal medicine, it has been utilized for stroke treatments, headache, wound healing, neurological disorder, etc. However, the understanding of its chemical matrices is still insufficient. Therefore, an online two-dimensional reversed phase liquid chromatography x hydrophilic interaction liquid chromatography (RPLCxHILIC) system coupled with mass spectrometry was applied for further detailed investigation of the chemical constituents in Buddleja dividii. In this two-dimensional liquid chromatography (2D-LC) method, a new atcolumn dilution (ACD) modulator was introduced in the 2D-LC system to solve the incompatibility problem of the mobile phase between two dimensions, which resulted in a 2D-LC analysis with high orthogonality. For the root extract, as one of the analyzed samples, the optimization of the 1 D and 2 D gradients was carried out carefully. With this new modulator, much better peak separation and better peak shape were achieved compared to two-dimensional liquid chromatography system using a traditional standard (TS) modulator. With a similar approach, the other four parts of Buddleja davidii were well separated. Comparing the different analyzed parts, flowers and leaves showed the most complex profiles. MS and MS/MS data were obtained successfully, which demonstrated the potential of the proposed RPLCxHILIC-MS system in the constituents' analysis of herbal medicine. However, due to the lack of reported reference information, 24 compounds could be tentatively identified.

Comparison of existing strategies for keeping symmetrical peaks in on-line Hydrophilic Interaction Liquid Chromatography x Reversed-Phase Liquid Chromatography despite solvent strength mismatch

“…This is carried out with a make-up pump in the first case and the ASM-valve (marketed by Agilent) in the second case. It should be noted that dilution after the switching valve was also recently reported under the name "at-column dilution" [21,22]. In this study, we considered the critical case in which the same column internal diameters (i.e.…”

“…More recently, the so-called "At-Column Dilution" (ACD) was proposed. In this case, dilution is carried out between the valve and the 2 Dcolumn by using an additional pump to empty the sample loop while the 2 D-pump dilutes the transferred fractions [21,22]. Regardless of the method, very large volumes which are proportional to the desired dilution ratio, are injected in 2 D. Nevertheless, improvements in both peak shape and peak intensity can be expected from the substantial decrease in eluent strength, which promotes band focusing at the 2 D-column inlet.…”

Strategies to circumvent the solvent strength mismatch problem in online comprehensive two‐dimensional liquid chromatography

“…HILIC × RPLC Phenolic acids Make-up flow [58] HILIC × RPLC Phenolics Make-up flow [95][96][97] HILIC × RPLC Oligosaccharides Make-up flow [80] HILIC × RPLC Standard mix At-column dilution [41] RPLC × HILIC Red ginseng extract At-column dilution [41] RPLC × HILIC Herbal medicine At-column dilution [42] Organic-SEC × RPLC Polymers Active solvent modulation [98,99] RPLC × RPLC Monoclonal antibody digests Active solvent modulation [100] HILIC × RPLC Small molecule probes Active solvent modulation [44] HILIC × RPLC Monoclonal antibodies Active solvent modulation [45] Diluting with a weak solvent (trapping columns) SFC × RPLC Pharmaceuticals [101] SFC × RPLC Citrus oil extract [101] SFC × RPLC Fatty acids [102] SFC × RPLC Triacylglycerols [103] SFC × RPLC Carotenoids [104] HILIC × RPLC Oligonucleotides [54] HILIC × RPLC Surfactants [55] HILIC × RPLC Phenolics [59,105] HILIC × RPLC Licorice metabolites [28] HILIC × RPLC Phenolic acids [58] HILIC × RPLC Lipids [106] HILIC × RPLC Cocoa bean metabolites [107] HILIC × RPLC Histone proteoforms [56] HILIC × RPLC Ginsenosides [57,108] HILIC × RPLC Oligosaccharides [80] HILIC × RPLC Microalgae digests [109] RPLC × HILIC Ginseng extract [110] IEC × RPLC Peptides [50] IEC × RPLC Dye extracts [111] HDC × RPLC Nanoparticles [112] RPLC × RPLC Vacuum gas oil [113] RPLC × RPLC Flavonoids [105] RPLC × RPLC Anabolic steroids…”

“…The sample is then diluted by the 2 D-pump with a weak initial mobile phase. This method was used in both RPLC × HILIC and HILIC × RPLC by Chen et al [41,42] for the analysis of red ginseng extract and herbal medicine.…”

Advances in ultra‐high‐pressure and multi‐dimensional liquid chromatography instrumentation and workflows

“…The latter can modulate 1D sample fractions, which are afterward mixed with the 2D gradient flow before reaching the 2D column. This device allows to modulate the dilution factor by adjusting the transfer flow rate and composition independent of the operating conditions used during the 1D and 2D analysis 149–151 …”