Development of a mesoporous silica based solid-phase extraction and ultra-performance liquid chromatography-MS/MS method for quantifying lignans in Justicia procumbens.

Justicia procumbens is a food and medicine homologous variety, popularly used for making vegetable soups. In this study, a novel mesoporous silica was synthesized and used as the sorbent of SPE for the purification of lignans from J. procumbens. A laboratory-made SPE cartridge was packed with 100 mg of mesoporous silica, which was washed with 10% methanol and eluted using 0.8 mL acetonitrile after sample loading. Afterward, the extract was analyzed by ultra-performance liquid chromatography (UPLC) and MS/MS. All the lignans were efficiently separated in 6 min with the noise level in the range of 50-150 cps. 6'-Hydroxy justicidin B, 6'-hydroxy justicidin A, justicidin B, chinensinaphthol methyl ether, justicidin C, and neojusticdin A were identified to be the dominant molecular species in J. procumbens with contents of 0.065-0.37 mg/g in three tested sample batches from different geographic origins. In conclusion, the proposed mesoporous silica based SPE UPLC-MS/MS method is efficient in linearity (R2 = 0.9989-0.9996), sensitivity (LOD ≤0.13 µg/kg and LOQ ≤0.42 µg/kg), precision (RSDintra-day ≤3.12 and RSDinter-day ≤4.56), and recovery (83.42-96.11%, RSD ≤2.88%).

Determination of kaurenoic acid in rat plasma using UPLC-MS/MS and its application to a pharmacokinetic study.

Kaurenoic acid (KA), a kaurane diterpene found in several medicinal plants, is an active ingredient with potential anti-inflammatory, anticonvulsant, antibacterial and antitumor activities. In this work, an ultra-performance liquid chromatography-tandem mass spectrometry method (UPLC-MS/MS) was firstly developed and validated to quantify kaurenoic acid in rat plasma. Rhein was chosen as the internal standard (IS) and the plasma was processed with one-step acetonitrile protein precipitation; the chromatographic separation was achieved on a HSS T3 (2.1 × 50 mm, 1.8 µm) column with the mobile phase consisting of acetonitrile and water containing 0.1% formic acid via gradient elution. An electrospray ionization source was applied and operated in the negative ion and multiple reaction monitoring (MRM) modes. Kaurenoic acid and IS were quantified using the transitions of m/z 301.2→301.2 (pseudo MRM) and m/z 283.2 → 238.9, respectively. The calibration curves were linear over the range of 5∼ 100 ng/mL (R2 = 0.990). The lower limit of quantification (LLOQ) was 5 ng/mL. The intra- and inter- day precision (RSD) ranged from 3.0% to 11.4%. The matrix effect and extraction recovery were within acceptable limits. The validated method was successfully applied to the pharmacokinetic study of kaurenoic acid in rats after oral administration at three dosages.

Chemical Profiling of Lobelia chinensis with High-Performance Liquid Chromatography/Quadrupole Time-of-Flight Mass Spectrometry (HPLC/Q-TOF MS) Reveals Absence of Lobeline in the Herb.

Lobelia chinensis is a kind of herbal medicine widely distributed and used in Asia. The chemical components of this herb, however, have not been well studied until now. Lobeline, as an essential and famous bioactive compound in Lobelia genus, has been assumed to be present in L. chinensis. In order to ascertain its presence and, more importantly, proper use of this herb, chemical profiling this herb with highly sensitive and high-resolution analytical mass spectrometry was applied. In this study, high-performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry (HPLC/Q-TOF MS) method was employed to systematically profile the chemical constituents of L. chinensis for the first time. Comparative chemical profiling study of L. chinensis and Lobelia inflata was also conducted to provide evidence whether lobeline is present or not. Piperidine alkaloids except for lobeline, alkaloid-lignan hybrids, flavonoids, polyacetylenes, nonanedioic acid, and some new phytochemicals were successfully identified in L. chinensis simultaneously. Comparing to the chemical profiles of L. inflata, lobeline was found to be absent in L. chinensis. All of the secondary metabolites in L. chinensis were determined with the HPLC/Q-TOF MS method. The absence of lobeline in L. chinensis was confirmed after this extensive study.

PRiME pass-through purification of lignans in Silybum marianum and UPLC-MS/MS analysis.

A PRiME (process, robustness, improvements, matrix effects, ease of use) pass-through cleanup procedure was developed for the extraction and purification of silychristins A and B, silybins A and B, isosilybins A and B, and silydianin in Silybum marianum. After optimizing the extracting solvent types and the sample loading volume, the crude extract was diluted to 3 mL with 95% acetonitrile and then loaded on the PRiME cartridge. The eluate was analyzed by ultra-performance liquid chromatography and tandem mass spectrometry (UPLC-MS/MS). All the target analytes were deprotonated as [M-H]- at m/z 481 by conducting collision-induced dissociation (CID), and the major fragment ions were m/z 463 ([M-H2O-H]-), 453 ([M-CO-H]-), 355 ([M-C6H6O3-H]-), 301 ([M355-CO2-H]-), and 179 ([C10H11O3]-). Afterwards, this method was validated in terms of linearity (R2 > 0.9990), intra-day precision (1.02%-3.79%), inter-day precision (1.59%-4.87%), sensitivity (LOD ≤ 0.45 µg·kg-1 and LOQ ≤ 1.50 µg·kg-1), and recovery (76.9-103.4%, RSD < 8.90%). Finally, the proposed protocol was successfully applied to eight batches of S. marianum samples. The total content of the seven active compounds varied amongst the batches from different places of origin.

Identification and Characterization of Kukoamine Metabolites by Multiple Ion Monitoring Triggered Enhanced Product Ion Scan Method with a Triple-Quadruple Linear Ion Trap Mass Spectrometer.

Kukoamines are a series of bioactive phytochemicals conjugated by a polyamine backbone and phenolic moieties. Understanding the structural diversity of kukoamine metabolites in plants is meaningful for drug discovery. In this study, an LC-MS/MS method was established for kukoamine profiling and characterization from lycii cortex (LyC) via a triple-quadrupole linear ion trap mass spectrometry (Q-TRAP). On the basis of the typical fragmentation of kukoamine, a diagnostic ion, which represents the features of the backbone and phenolic substitute, was chosen as the product ion for precursor ion scan, and then the screened precursor ions were applied to a successive multiple ion monitoring triggered enhanced product ion scan (MIM-EPI) to simultaneously present the profile survey and MS/MS acquisition. Because the MIM narrowed the ion scan range in Q1 and the ion trap enhanced the ion fragments passing through Q2, the qualitative capability of quadrupole MS can be greatly improved, especially for capture of the uncommon metabolites. There are 12 kukoamine metabolites identified from LyC, with either spermine or spermidine backbone and with conjugation of one to three dihydrocaffeoyls or other kinds of phenolic moieties. Except for kukoamines A and B, other metabolites were identified in LyC for the first time. This approach can be utilized for metabolite identification in other substrates.

Direct analysis of herbal powders by pipette-tip electrospray ionization mass spectrometry.

Conventional electrospray ionization mass spectrometry (ESI-MS) is widely used for analysis of solution samples. The development of solid-substrate ESI-MS allows direct ionization analysis of bulky solid samples. In this study, we developed pipette-tip ESI-MS, a technique that combines pipette tips with syringe and syringe pump, for direct analysis of herbal powders, another common form of samples. We demonstrated that various herbal powder samples, including herbal medicines and food samples, could be readily online extracted and analyzed using this technique. Various powder samples, such as Rhizoma coptidis, lotus plumule, great burdock achene, black pepper, Panax ginseng, roasted coffee beans, Fructus Schisandrae Chinensis and Fructus Schisandrae Sphenantherae, were analyzed using pipette-tip ESI-MS and quality mass spectra with stable and durable signals could be obtained. Both positive and negative ion modes were attempted and various compounds including amino acids, oligosaccharides, glycosides, alkaloids, organic acids, ginosensides, flavonoids and lignans could be detected. Principal component analysis (PCA) based on the acquired mass spectra allowed rapid differentiation of closely related herbal species.