Drug Metabolite Cluster-Based Data-Mining Method for Comprehensive Metabolism Study of 5-hydroxy-6,7,3',4'-tetramethoxyflavone in Rats.

The screening of drug metabolites in biological matrixes and structural characterization based on product ion spectra is among the most important, but also the most challenging due to the significant interferences from endogenous species. Traditionally, metabolite detection is accomplished primarily on the basis of predicted molecular masses or fragmentation patterns of prototype drug metabolites using ultra-high performance liquid chromatography coupled with high-resolution mass spectrometry (UHPLC-HRMS). Although classical techniques are well-suited for achieving the partial characterization of prototype drug metabolites, there is a pressing need for a strategy to enable comprehensive drug metabolism depiction. Therefore, we present drug metabolite clusters (DMCs), different from, but complementary to, traditional approaches for mining the information regarding drugs and their metabolites on the basis of raw, processed, or identified tandem mass spectrometry (MS/MS) data. In this paper, we describe a DMC-based data-mining method for the metabolite identification of 5-hydroxy-6,7,3',4'-tetramethoxyflavone (HTF), a typical hydroxylated-polymethoxyflavonoid (OH-PMF), which addressed the challenge of creating a thorough metabolic profile. Consequently, eight primary metabolism clusters, sixteen secondary metabolism clusters, and five tertiary metabolism clusters were proposed and 106 metabolites (19 potential metabolites included) were detected and identified positively and tentatively. These metabolites were presumed to generate through oxidation (mono-oxidation, di-oxidation), methylation, demethylation, methoxylation, glucuronidation, sulfation, ring cleavage, and their composite reactions. In conclusion, our study expounded drug metabolites in rats and provided a reference for further research on therapeutic material basis and the mechanism of drugs.

Chemical Constituent Profiling of Paecilomyces cicadae Liquid Fermentation for Astragli Radix.

Astragli Radix (AR) is one of the most popular traditional Chinese medicines with chemical constituents including flavonoids and saponins. As recently evidenced, some fungi or their fermentation liquid may have the potential to affect the bioactive constituents and different pharmacological effects of AR. Thus, the composition of fermented AR (FAR) produced by Paecilomyces cicadae (Miquel) Samson in liquid-state fermentation was investigated using a UHPLC-LTQ-Orbitrap mass spectrometer in both positive and negative ion modes. Firstly, the MSn data sets were obtained based on a data-dependent acquisition method and a full scan-parent ions list-dynamic exclusion (FS-PIL-DE) strategy. Then, diagnostic product ions (DPIs) and neutral loss fragments (NLFs) were proposed for better constituent detection and structural characterization. Consequently, 107 constituents in total, particularly microconstituents in FAR and AR, were characterized and compared in parallel on the same LTQ-Orbitrap instrument. Our results indicated that AR fermentation with Paecilomyces significantly influenced the production of saponins and flavonoids, especially increasing the content of astragaloside IV. In conclusion, this research was not only the first to show changes in the chemical components of unfermented AR and FAR, but it also provides a foundation for further studies on the chemical interaction between microbiota and AR.

Profiling and identification of chlorogenic acid metabolites in rats by ultra-high-performance liquid chromatography coupled with linear ion trap-Orbitrap mass spectrometer.

1. Chlorogenic acids (CGAs), one kind of major bioactive constituents isolated from Flos Lonicera Japonica, possess many biological activities, such as antibacterial, antioxidant and antiviral activities. In this study, we established an efficient strategy using ultra-high-performance liquid chromatography coupled with linear ion trap-Orbitrap mass spectrometry (UHPLC-LTQ-Orbitrap MS) to profile the in vivo metabolic fate of CGAs in rat urine and plasma. 2. The extract from Flos Lonicera Japonica was orally administrated to Sprague-Dawley (SD) rats at a dose of 1000 mg/kg body weight. Then, a combination of various post-acquisition data mining methods, including high-resolution extracted ion chromatogram (HREIC) and multiple mass defect filters (MMDFs) and diagnostic product ions (DPIs), were adopted to characterize the known and unknown CGA metabolites in SD rats. 3. As a result, a total of 68 CGA metabolites were unambiguously or tentatively screened and characterized. These metabolites, including 18 prototype compounds and 50 metabolites, were deduced to be yielded via methylation, hydrogenation, demethylation, dehydration, sulfate conjugation, glucuronide conjugation, glycosylation conjugation and their composite reactions, which mainly occurred to caffeoylquinic acids, dicaffeoylquinic acids, p-coumaroylquinic acids and feruloylquinic acids. 4. In conclusion, this study profiled CGA metabolites, which are useful in understanding the in vivo metabolic fate, effective forms, and pharmacological and toxic actions of CGAs.

[Identification of metabolites of Danshensu in vivo in rats].

To identify the metabolites of Danshensu in plasma and urine in rats by using UHPLC-LTQ-Orbitrap method. After oral gavage of Danshensu CMC-Na suspension in SD rats, urine and plasma samples were collected and processed by solid phase extraction. ACQUITY UPLC BEH C18 column (2.1 mm×100 mm, 1.7 µm) was utilized, with 0.1% formic acid (A)-acetonitrile (B) solution as the mobile phase for gradient elution. Negative electrospray ion mode based data-acquisition method was established to collect the mass spectrometry data of biological samples. As a result, Danshensu and 21 Danshensu Ⅰ phase and Ⅱ phase metabolites were finally identified according to the accurate mass measurements, mass fragmentation behaviors and comparing with the reference standards. The main metabolic pathways included dehydration, methylation, glucuronide conjugation, sulfate conjugation and their composite reactions. Consequently, our study expounded metabolites of Danshensu in rats based on UHPLC-LTQ-Orbitrap method and provided a reference for further researches on therapeutic material basis and mechanism of Danshensu.

The chemical transformations for Radix Astragali via different alkaline wash conditions by quantitative and qualitative analyses.

Radix Astragali is a famous Chinese traditional and folk medicine with a wide range of medicinal values in clinic. In this study, an analytical efficient strategy based on UHPLC-QQQ-MS/MS and UHPLC-LTQ-Orbitrap-MS/MS was established to explore and reveal the chemical transformations for Radix Astragali under different alkaline wash conditions for analytical sample preparation. Firstly, a rapid and sensitive UHPLC-QQQ-MS/MS method for the quantification of 14 main constituents in Radix Astragali has been developed and validated. Secondly, according to the standard substance comparison, accurate mass measurements, mass fragmentation behaviors and related literatures, a total of 102 components have been screened and identified using UHPLC-LTQ-Orbitrap method. Among them, 47 compounds are saponins, and the other 55 are flavonoids. Consequently, there were two chemical transformations including hydrolysis and degradation observed when Radix Astragali was treated with alkali. Besides, hydrolysis of glycosides and acetyl played a considerably important role in the process of sample preparation. It has been proved that 10 % ammonia could relatively guarantee the high content of astragaloside IV and avoid the over-degradation of most chemical ingredients in Radix Astragali. In conclusion, this work would provide a scientific and practical method for quality control of Radix Astragali as well as its compound preparations.

Comprehensive analysis of the chemical constituents in sulfur-fumigated Lonicerae Japonicae Flos using UHPLC-LTQ-Orbitrap mass spectrometry.

Lonicerae Japonicae Flos (LJF), a kind of traditional Chinese medicines (TCMs), has functions of detoxifying and evacuating heat. In the study, a method based on ultra-high performance liquid chromatography coupled with linear ion trap-Orbitrap mass spectrometry (UHPLC-LTQ-Orbitrap MS) was developed for the chemical constituent analysis of organic acids, flavonoids, iridoids and new-generated compounds in sulfur-fumigated LJF (SF-LJF). Based on the accurate mass measurement (< ± 5 ppm), chromatographic behavior and diagnostic product ions (DPIs), 113 constituents were unambiguously or tentatively characterized from SF-LJF extract, including 46 chlorogenic acids, 19 flavonoids, 29 iridoid glycosides and 19 newly-generated compounds (including 17 sulfur-containing derivatives). In addition, 5-CQA (5-caffeoylquinic acid, chlorogenic acid) was chosen to be sulfur-fumigated for the result validation. It was found that the most significant change of LJF after sulfur fumigation was the occurrence of sulfate or sulfite esterification reactions, which resulted in the emergence of many new sulfur-containing components. Our results demonstrated that the established method was a useful and efficient analytical tool to comprehensively characterize the material basis of SF-LJF, and also an excellent guidance of quality control about LJF.

Comprehensive metabolism study of polydatin in rat plasma and urine using ultra-high performance liquid chromatography coupled with high-resolution mass spectrometry.

Polydatin is one of the main bioactive constituents isolated from Polygonum cuspidatum Sieb. et Zucc., which possesses various pharmacological activities. In this study, we established an efficient strategy based on ultra-high performance liquid chromatography coupled with high-resolution mass spectrometry to uncover polydatin metabolites in rat urine and plasma. Firstly, multiple mass defect filters (MMDFs) combined with high-resolution extracted ion chromatograms (HREICs) was utilized to perform post-acquisition data-mining in ESI-MS1 stage. Secondly, metabolite candidates of polydatin were expounded systematically on the basis of diagnostic product ions (DPIs), chromatographic retention times, accurate mass, and neutral loss fragments (NLFs). Consequently, a total of 41 metabolites (polydatin included) were detected and identified tentatively in 12 min. These metabolites, including 40 in rat urine and 7 in rat plasma, were presumed to generate through glucuronidation, sulfation, deglucosylation, dehydrogenation, methylation, hydrogenation, hydroxylation and their composite reactions. Meanwhile, metabolite clusters, which were set in the form radially, were found in this study. In conclusion, our study expounded polydatin metabolites in rats and provided a reference for further researches on therapeutic material basis and mechanism of polydatin.

Report from the BIT's 1st Annual World Congress of Biomedical Engineering Held in Xi'an, China, 9-11 November 2017.

We are delighted to present within this meeting report the abstracts of the "BIT's 1st World Congress of Biomedical Engineering 2017" which has been hold in Xi'an in China [...].

Report from the BIT's 4th Annual World Congress of High-Tech Acupuncture and Integrative Medicine Held in Xi'an, China, 25-27 September 2017.

High-tech acupuncture is an example of a very successful cross-over between tradition and innovation. "The Annual World Congress of High-Tech Acupuncture and Integrative Medicine" was successfully organized on four occasions, and attracted more than 800 participants from 55 countries [...].

Analysis of multiplex PCR fragments with PMMA microchip.

Microchip electrophoresis is a promising technique for analysis of bio-molecules. It has the advantages of fast analysis, high sensitivity, high resolution and low-cost of samples. Plastic chip has the potential of mass production for clinical use for its advantages in biocompatibility and low cost. In this work, the method for fabrication of poly(methyl methacrylate) (PMMA) chip was described, and conditions for DNA separation were investigated with the chip. The PMMA microchip was used for detection of multiplex PCR products of 18 and 36 cases with SARS and hepatitis B virus infection under optimized separation conditions. Microchip electrophoresis showed higher sensitivity, higher resolution and less time consumption when compared with gel electrophoresis. The microchip electrophoresis with PMMA chip provided a rapid, sensitive and reliable method for analysis of multiplex PCR products.