Ying Xue

Herbal medicine has been widely valued because of its remarkable efficacy and minimal side effects. The quantitative analysis of herbal medicines is essential to ensure their safety and efficacy. The simultaneous detection of multiple quality markers (Q-markers) has emerged as an important approach and trend in herbal medicine quality control. In recent years, non-targeted screening has become an effective strategy for the discovery and identification of unknown compounds. This study developed a non-targeted screening and quantitative analysis strategy to discover, identify and quantify the multiple components that truly represent the efficacy of Wuling capsule. Within this strategy, 18 types of flavonoids were tentatively discovered and identified from Wuling capsule by analyzing mass cleavage pathways, the precise molecular weights of compounds, and comparing the data with a database. Ten types of flavonoids were determined after the comparison of the standards. Additionally, following the evaluation of the regression equation, linear range, limit of detection (LOD), limit of quantitation (LOQ), precision, repeatability, and recovery of the proposed quantitative method, six flavonoids were quantified. This method successfully screened, identified, and quantified the potential active components in Wuling capsule, providing insights for improving the quality control standards in other herbal medicines.

Nucleosides can be used as quality evaluation indicators of Tricholoma matsutake. In this work, a new ultra-performance liquid chromatography-tandem mass spectrometry (UPLC/MS) strategy for quantitative analysis of multiple components using a single marker (QAMS) was proposed to determine nine nucleosides (adenosine, cytidine, guanosine, inosine, uridine, 2-deoxyadenosine, 2-deoxycytidine, 2-deoxyguanosine, and 2-deoxyuridine) in T. matsutake. Guanosine was set as the internal reference substance, whose content in T. matsutake was determined using the conventional external standard method. Relative correction factors (RCFs) between guanosine and eight other nucleosides were measured. The concentrations of the eight components were calculated with the obtained RCFs by QAMS. An ultrasonic extraction method is used for sample preparation. This method was validated to be sensitive, precise, and accurate with the LODs of 0.311.9ng, the overall intraday and interday variations less than 4.08, and the overall recovery over 89.0. The correlation coefficients (r²) of the calibration curves were higher than 0.9918. The values of vector angle analysis were above 0.99845, which indicates no significant differences between the conventional external standard method and the present QAMS method. As far as we know, this is also the first report of UPLC/MS analysis of nucleoside compounds by QAMS, providing an efficient and feasible quality assessment method for other natural products containing nucleosides.

Human beta-defensins (hBDs) play an important role in the host defense against various microbes, showing different levels of antibacterial activity and salt resistance in vitro. It is of interest to investigate whether can chimeric hBD analogs enhanced antibacterial activity and salt resistance. In this study, we designed a chimeric human defensin, named H4, by combining sequences of human beta-defensin-3 (hBD-3) and human beta-defensin-4 (hBD-4), then evaluated its antibacterial activity, salt resistance, and cytotoxic effects. The result showed that the antibacterial activity of H4 against most tested strains, including Klebsiella pneumonia, Enterococcus faecalis, Staphyloccocus aureus, Escherichia coli, Pseudomonas aeruginosa, Klebsiella pneumonia, and Acinetobacter baumannii was significantly improved compared to that of hBD-3 and hBD-4. Notably, H4 exhibited significantly better antibacterial activity against multidrug resistant isolate A. baumannii MDR-ZJ06 than commonly used antibiotics. Chimeric H4 still showed more than 80% antibacterial activity at high salt concentration (150 μM), which proves its good salt tolerance. The cytotoxic effect assay showed that the toxicity of H4 to Hela, Vero, A549 cells and erythrocytes at a low dose (<10 μg/ml) was similar to that of hBD-3 and hBD-4. In conclusion, given its broad spectrum of antibacterial activity and high salt resistance, chimeric H4 could serve as a promising template for new therapeutic antimicrobial agents.

In this work, a new quantitative analysis method of multi‐components analysis via a single marker strategy coupled with high‐performance liquid chromatography (HPLC) analysis, was proposed to analyze nine nucleosides (cytidine, uridine, 2′‐deoxyuridine, inosine, guanosine, 2′‐deoxyguanosine, thymidine, adenosine, and 2′‐deoxyadenosine) as quality control markers in Rhizoma Paridis. Guanosine was set as the internal reference substance, whose content in Rhizoma Paridis was determined using conventional external standard method. Then, relative correction factors between guanosine and the other eight nucleosides were measured respectively. The amounts of the other eight components were calculated according to the relative correction factors by the quantitative analysis of multi‐components via a single marker method. Finally, the result of vector angle cosine analysis showed that there was no significant difference of the contents between the external standard method and the quantitative analysis of multi‐components via a single marker method, indicating that the quantitative analysis of multi‐components via a single marker method can be applied for the quality control of Rhizoma Paridis. As far as we know, this is also the first report to analyze nucleosides by the quantitative analysis of multi‐components via a single marker method, providing an efficient and promising quality assessment method for other traditional Chinese medicine containing nucleosides.

So far, there is no report on the quality evaluation of lemonade available in the market. In this study, a sample preparation method was developed for the determination of flavonoid glycosides by ultra-performance liquid chromatography–mass spectrometry (UPLC-MS) based on vortex-assisted dispersive liquid-liquid microextraction. First, potential flavonoids in lemonade were scanned and identified by ultra-performance liquid chromatography–time of flight mass spectrometry (UPLC-TOF/MS). Five flavonoid glycosides were identified as eriocitrin, narirutin, hesperidin, rutin, and diosmin according to the molecular formula provided by TOF/MS and subsequent confirmation of the authentic standard. Then, an ultra-performance liquid chromatography–triple quadrupole mass spectrometry (UPLC-QqQ/MS) method was developed to determine these five flavonoid glycosides in lemonade. The results showed that the content of rutin in some lemonade was unreasonably high. We suspected that many illegal manufacturers achieved the goal of low-cost counterfeiting lemonade by adding rutin. This suggested that it was necessary for relevant departments of the state to make stricter regulations on the quality standards of lemonade beverages.

So far, there has been no quality evaluation of Tricholoma matsutake. Nucleic acid compounds are a kind of functional ingredient in T. matsutake that is beneficial to human health. In this study, a UPLC-TOF/MS method was first used to scan and identify the potential nucleic acid compounds in T. matsutake. Based on the calculation of the molecular formula and subsequent confirmation by authentic standards, 15 nucleic acid compounds were unambiguously identified: adenosine, cytidine, guanosine, inosine, thymidine, uridine, xanthosine dehydrate, 2′-deoxyadenosine, 2′-deoxycytidine, 2′-deoxyguanosine, 2′-deoxyuridine, adenosine 5′-monophosphate, cytidine 5′-monophosphate, guanosine 5′-monophosphate, and uridine 5′-monophosphate. Then, a UPLC-QqQ/MS method was developed for the subsequent quantitative analysis. After validating the limits of quantification, detection, precision, repeatability, and recovery through a calibration curve, the content of 15 nucleic acid compounds was determined by the proposed UPLC-QqQ/MS method in 80 T. matsutake samples collected from different regions in Sichuan province, Southwest China. After the statistical analysis, we suggest that the total content of nucleic acid compounds in the qualified T. matsutake should be higher than 24.49 mg/100 g. The results indicated that the combined use of UPLC-TOF/MS and UPLC-QqQ/MS is efficient for fast identification and determination of nucleic acid compounds to comprehensively evaluate the quality of T. matsutake.

In the present study, 11 non-anthocyanin polyphenols, gallic acid, protocatechuate, vanillic acid, syringic acid, ferulic acid, quercetin, catechin, epicatechin, epigallocatechin gallate, gallocatechin gallate and epicatechin gallate—were firstly screened and identified from blueberries using an ultra performance liquid chromatography–time of flight mass spectrography (UPLC-TOF/MS) method. Then, a sample preparation method was developed based on vortex-assisted dispersive liquid-liquid microextraction. The microextraction conditions, including the amount of ethyl acetate, the amount of acetonitrile and the solution pH, were optimized through the multi-objective response surface methodology and desirability function approach. Finally, an ultra performance liquid chromatography–triple quadrupole mass spectrography (UPLC-QqQ/MS) method was developed to determine the 11 non-anthocyanin polyphenols in 25 commercial blueberry samples from Sichuan province and Chongqing city. The results show that this new method with high accuracy, good precision and simple operation characteristics, can be used to determine non-anthocyanin polyphenols in blueberries and is expected to be used in the analysis of other fruits and vegetables.

The peels of citrus fruits (Pericarpium Citri Reticulatae, PCR) have long been used in traditional Chinese medicines (TCMs). Polymethoxylated flavonoids (PMFs) were found to be the main components present in PCR extracts, but their metabolism remains unclear which restrain the utilization of this TCM. In the present work, rat liver microsomes were immobilized on magnetic nanoparticles (LMMNPs) for in vitro metabolic study on the whole PMFs of PCR. LMMNPs were characterized by transmission electron microscope and Fourier-transform infrared spectrum. The relative enzyme binding capacity of LMMNPs was estimated to be about 428 μg/mg from thermogravimetric analysis. Incubation of LMMNPs with PMFs produced demethylated metabolites of PMFs, six of which were identified by ultrahigh pressure liquid chromatography–mass spectrometry (UPLC–MS/MS). The 3′-hydroxylated tangeretin (T3) was detected from the metabolites of tangeretin for the first time, which suggested that 4′-demethylated and 3′-hydroxylated derivative of tangeretin (3′-hydroxy-5,6,7,8,4′-pentamethoxyflavone, T4) was not only derived from 4′-demethylated tangeretin (T2) as previously reported, but also from T3. This is the first investigation of the metabolism of the whole PMFs, which may shed light on the mechanism of action of PCR.