Comparison of the chemical profiles of fresh‐raw and dry‐processed Juglans mandshurica

The processing of Juglans mandshurica Maxim. is important to reduce its toxicity and enhance its efficacy. Simple, efficient, and sensitive ultra‐high performance liquid chromatography coupled with a time‐of‐flight mass spectrometry based chemical profiling approach was proposed to rapidly evaluate the chemical difference between fresh and dry samples. Under the optimized ultra‐high performance liquid chromatography and quadrupole time‐of‐flight tandem mass spectrometry conditions, 81 significantly different compounds were rapidly discovered using principal component analysis, and then tentatively identified by comparison with reference substances or inferred through mass spectral fragment ion analysis and literature data. These compounds included 35 naphthoquinones, 11 diarylheptanoids, nine flavonoids, eight triterpenes, 12 phenolic acids, and six aliphatics. The results demonstrated that chemical reactions occurring during processing could be used to elucidate the processing mechanism of Juglans mandshurica Maxim. This study provides a novel approach to identifying complicated components of various complex mixtures in fresh‐raw and dry‐processed traditional Chinese medicines, which could be used as a valid analytical method to further understand the processing mechanisms of these medicines, as well as providing intrinsic quality control of the medicines and their processed products.     

The comparative research on constituents of Radix Aconiti and its processing by HPLC quadrupole TOF-MS

Based upon the regulations stipulated by the State Food and Drug Administration of China, only the processed, detoxified tubers and roots of Aconitum are allowed to be administered orally, used in clinical decoctions and adopted as raw materials for pharmaceutical manufacturing, so the processing principle of preparation of Radix Aconiti is important for ensuring the Radix Aconiti praeparata quality. A simple approach was described for HPLC‐Q‐TOF‐MS screening and identification of many of the aconitine alkaloids present in unprocessed Radix Aconiti and Radix Aconiti praeparata. To compare their fingerprints, the processing principle of preparation of Radix Aconiti was developed. Twenty‐nine compounds and 26 compounds were assigned to aconitine alkaloids and tentatively identified by comparing accurate mass and fragments information with that of the authentic standards or by mass spectrometry analysis and retrieving the reference literature. The nonester alkaloids were almost the same. The diester diterpene alkaloids were decreased, the monoester‐diterpene alkaloids were increased and lipo‐alkaloids decreased obviously in the processing of the preparation. These transformed components could be regarded as potential chemical markers that can be used to distinguish between raw and processed herbs. Copyright © 2012 John Wiley & Sons, Ltd.     

Determination of the chemical constituents of the different processed products of Anemarrhena asphodeloides Rhizomes by high‐performance liquid chromatography quadrupole time‐of‐flight mass spectrometry

In this work, high‐performance liquid chromatography (HPLC) coupled with a hybrid quadrupole time of‐flight mass spectrometry (Q‐TOF‐MS/MS) was used to study chemical compositions of different processed products of Rhizoma Anemarrhenae (RA). A Grace Alltima^TM C₁₈ column (250 × 4.6 mm, 5 µm) was used for separation. Mobile phase consisted of 0.1% formic acid and acetonitrile, using gradient elution. ESI‐MS data was acquired in both positive and negative mode. The experiment was established on the basis of a series of reference substances (two xanthone and seven saponins) to qualitatively identify the chemical compounds of different processed products of RA by MS analysis. There was no difference in the type of chemical constituents between different processed products of RA. A total of 25 compounds were identified, including four xanthones, 21 steroidal saponins and eight pairs of isomers. Copyright © 2015 John Wiley & Sons, Ltd.     

Chemical differentiation of volatile compounds in crude and processed Atractylodis Macrocephalae Rhizoma by using comprehensive two‐dimensional gas chromatography with time‐of‐flight mass spectrometry combined with multivariate data analysis

In the present study, comprehensive 2D GC—TOF‐MS combined with multivariate data analysis was applied to analyze the differences of the volatile components in crude and processed Atractylodis Macrocephalae Rhizoma (AMR) samples. As a result, 26 compounds that were found in crude AMR samples disappeared in processed AMR samples, and 19 compounds were newly generated and identified in AMR after processing with wheat bran. Meanwhile, principal component analysis demonstrated that there were significant chemical differences between crude and processed AMR samples, and processing procedure caused obvious quantitative and qualitative changes of volatile components in AMR. The established method could be used to explain the chemical differentiation between crude and processed AMR, and to further understand the processing mechanism of herbal medicines.     

Qualitative and quantitative analysis of the chemical constituents in Mahuang‐Fuzi‐Xixin decoction based on high performance liquid chromatography combined with time‐of‐flight mass spectrometry and triple quadrupole mass spectrometers

High‐performance liquid chromatography coupled with time‐of‐flight mass spectrometry (HPLC‐TOF/MS) and high‐performance liquid chromatography–triple quadrupole mass spectrometry (HPLC‐QQQ/MS/MS) were utilized to clarify the chemical constituents of Mahuang‐Fuzi‐Xixin Decoction. There are 52 compounds, including alkaloids, amino acids and organic acids were identified or tentatively characterized by their characteristic high resolution mass data by HPLC‐QQQ/MS/MS. In the subsequent quantitative analysis, 10 constituents, including methyl ephedrine, aconine, songrine, fuziline, neoline, talatisamine, chasmanine, benzoylmesaconine, benzoylaconine and benzoylhypaconine were simultaneously determined by HPLC‐QQQ/MS/MS with multiple reaction monitoring mode. Satisfactory linearity was achieved with wide linear range and fine determination coefficient (r > 0.9992). The relative standard deviations (RSD) of inter‐ and intra‐day precisions were <3%. This method was also validated by repeatability, stability and recovery with RSD <3% respectively. A highly sensitive and efficient method was established for chemical constituents studying, including identification and quantification of Mahuang‐Fuzi‐Xixin decoction.     

Development and validation of a UHPLC‐MS/MS bioanalytical method to quantify in plasma the analgesic candidate PT‐31 following a preliminary pharmacokinetic study in rats

A selective and sensitive UHPLC‐MS/MS bioanalytical method to determine PT‐31, an analgesic drug candidate, in rat plasma was developed and validated. Analyses were performed using a UHPLC‐MS/MS system equipped with an electrospray ionization interface operating in the positive ionization mode using a C₁₈ reversed‐phase column with a mobile phase of water:acetonitrile (68:31, v/v) containing 0.1% acetic acid eluting in a gradient mode with a flow rate of 0.3 mL/min. Plasma samples were deproteinized with cold acetonitrile containing 0.01% TFA (1:2, v/v) and 50 μL of the supernatant were injected into the system. PT‐31 and phenytoin (internal standard) retention times were roughly 1.0 and 1.5 min, respectively. Linear standard curves were plotted for the 0.01–10 µg/mL concentration range, with a coefficient of determination > 0.99. The method's precision was over 88%. Maximum intra‐ and inter‐day relative standard deviations were 14.6% and 11.6%, respectively. Interfering substances were not detected in the chromatogram, indicating that the method was specific. PT‐31 stability was assessed under different temperature and storage settings. The method was used to characterize PT‐31 plasma pharmacokinetics following administration of 5 mg/kg i.v. to Wistar rats. Therefore, the method described is sensitive, linear, precise and specific enough to determine PT‐31 in preclinical pharmacokinetic investigations. Copyright © 2015 John Wiley & Sons, Ltd.     

Identifying the chemical markers in raw and wine-processed Scutellaria baicalensis by ultra-performance liquid chromatography/quadrupole time-of-flight mass spectrometry coupled with multiple statistical strategies

Herb processing is a typical pharmaceutical preparation process for traditional Chinese medicine. After processing, its clinical applications and pharmacological effects vary greatly, which is most commonly attributed to the changing chemical properties between raw herb and processed products. In this work, a total of 53 chemical compounds were detected, among which 17 compounds were identified as discriminatory chemicals between raw and wine-processed Scutellaria baicalensis, and 10 components were identified as chemical markers with a cumulative content contribution of 88.75%. In addition, this work revealed that the best wine-processed time was 18 min by investigating the changes of chemical markers in S. baicalensis during processing. This work demonstrated that ultra-performance liquid chromatography/quadrupole time-of-flight mass spectrometry coupled with multiple statistical strategies is an effective approach for screening and identifying discriminatory chemical markers in complex traditional Chinese medicine.     

Characterization and discrimination of raw and vinegar‐baked Bupleuri radix based on UHPLC‐Q‐TOF‐MS coupled with multivariate statistical analysis

Bupleuri Radix is a commonly used herb in clinic, and raw and vinegar‐baked Bupleuri Radix are both documented in the Pharmacopoeia of People's Republic of China. According to the theories of traditional Chinese medicine, Bupleuri Radix possesses different therapeutic effects before and after processing. However, the chemical mechanism of this processing is still unknown. In this study, ultra‐high‐performance liquid chromatography with quadruple time‐of‐flight mass spectrometry coupled with multivariate statistical analysis including principal component analysis and orthogonal partial least square‐discriminant analysis was developed to holistically compare the difference between raw and vinegar‐baked Bupleuri Radix for the first time. As a result, 50 peaks in raw and processed Bupleuri Radix were detected, respectively, and a total of 49 peak chemical compounds were identified. Saikosaponin a, saikosaponin d, saikosaponin b₃, saikosaponin e, saikosaponin c, saikosaponin b₂, saikosaponin b₁, 4′′‐O‐acetyl‐saikosaponin d, hyperoside and 3′,4′‐dimethoxy quercetin were explored as potential markers of raw and vinegar‐baked Bupleuri Radix. This study has been successfully applied for global analysis of raw and vinegar‐processed samples. Furthermore, the underlying hepatoprotective mechanism of Bupleuri Radix was predicted, which was related to the changes of chemical profiling.     

Global detection and identification of components from crude and processed traditional Chinese medicine by liquid chromatography connected with hybrid ion trap and time‐of‐flight‐mass spectrometry

We herein present a chemical profiling method to efficiently process the information acquired by ultra fast liquid chromatography (UFLC)‐electrospray ionization source in combination with hybrid ion trap and high‐resolution time‐of‐flight mass spectrometry (UFLC‐(ESI)‐IT‐TOF/MS), facilitating the structural determination of serial components contained in crude or processed traditional Chinese medicine (TCM). Under the optimized UFLC and IT‐TOF‐MSⁿ conditions, over 39 compounds were separated and detected in crude or processed Fructus corni within 25 min. The components were identified by comparing the mass spectra and retention time with reference compounds, or tentatively assigned by elucidating low‐energy collision‐induced dissociation (CID) fragment ions and matching empirical molecular formula with that of the published compounds. Several factors in the processing procedure were examined. The experimental results demonstrate that the chemical reactions that occurred in the processing procedure can be used to elucidate the processed mechanism of F. corni, which is regularly affected by the processing conditions. This study provides a novel approach and methodology to identify the complicated components from various complex mixtures such as crude TCM, processed TCM, and biological samples. It can be used as a valid analytical method for further understanding the processing mechanism of TCM, along with the intrinsic quality control of TCM and its processed product.     

Joint MS‐based platforms for comprehensive comparison of rat plasma and serum metabolic profiling

Metabolomics is a rapidly growing field in the comprehensive understanding of cellular and organism‐specific responses associated with perturbations induced by medicines, chemicals and environment. Blood matrices are frequently used in clinical and biological studies. In this study, we compared metabolic profiling between rat plasma and serum using complementary platforms of gas chromatography–mass spectrometry (GC‐MS) and liquid chromatography–quadruple time‐of‐flight–mass spectrometry (LC‐QTOF‐MS). The sample types that were tested included plasma prepared with K₂EDTA and serum collected using venous blood collection protocols. The results of peak area variation for each detected metabolite/feature in the quality control samples showed a good reproducibility in LC‐QTOF‐MS and better reproducibility in GC‐MS. In GC‐MS analysis: (a) 25.8% of the defined metabolites differed serum from plasma profiling (t‐test, p < 0.05); and (b) serum possessed higher sensitivity than plasma for its generally higher peak intensity in the metabolic profiling. In LC‐QTOF‐MS analysis, 13 (in positive ion mode) and seven (in negative ion mode) important metabolites were identified as mainly contributing to the separation between serum and plasma. Copyright © 2014 John Wiley & Sons, Ltd.     

Ultrasound/microwave‐assisted extraction and comparative analysis of bioactive/toxic indole alkaloids in different medicinal parts of Gelsemium elegans Benth by ultra‐high performance liquid chromatography with MS/MS

Indole alkaloids are the main bioactive/toxic components in Gelsemium elegans Benth. To determine the distribution and contents of indole alkaloids in its different medicinal parts, a novel and rapid method using ultra‐high performance LC (UPLC) with MS/MS has been established and validated with an optimized ultrasound/microwave‐assisted extraction method. Four constituents, namely, humantenidine, humantenmine, gelsemine, and koumine, were simultaneously determined in 6 min. Chromatographic separation was achieved on an ultra‐high performance LC BEH C₁₈ column with a gradient mobile phase consisting of methanol and water (containing 0.1% formic acid both in methanol and water) at a flow rate of 0.3 mL/min. The detection was performed on a triple quadrupole electrospray MS/MS by positive ion multiple‐reaction monitoring mode. All the analytes showed good linearity (r ≥ 0.9934) within a concentration range from 0.1–25 μg/mL with a LOQ of 25–50 ng/mL. The overall intra‐ and intervariations of four components were <4.7% with an accuracy of 97.3–101.3%. The analysis results showed that there were remarkable differences in the distribution and contents of four chemical markers in the roots, stems, and leaves of G. elegans Benth. The findings can provide necessary and meaningful information for the rational utilization of its resources.     

An isomer-specific high-energy collision-induced dissociation MS/MS database for forensic applications: a proof-of-concept on chemical warfare agent markers

Spectra database search has become the most popular technique for the identification of unknown chemicals, minimizing the need for authentic reference chemicals. In the present study, an isomer‐specific high‐energy collision‐induced dissociation (CID) MS/MS spectra database of 12 isomeric O‐hexyl methylphosphonic acids (degradation markers of nerve agents) was created. Phosphonate anions were produced by the electrospray ionization of phosphonic acids or negative‐ion chemical ionization of their fluorinated derivatives and were analysed in a hybrid magnetic‐sector–time‐of‐flight tandem mass spectrometer. A centre‐of‐mass energy (E_com) of 65 eV led to an optimal sequential carbon–carbon bond breakage, which was interpreted in terms of charge remote fragmentation. The proposed mechanism is discussed in comparison with the routinely used low‐energy CID MS/MS. Even‐mass (odd‐electron) charge remote fragmentation ion series were diagnostic of the O‐alkyl chain structure and can be used to interpret unknown spectra. Together with the odd‐mass ion series, they formed highly reproducible, isomer‐specific spectra that gave significantly higher database matches and probability factors (by 1.5 times) than did the EI MS spectra of the trimethylsilyl derivatives of the same isomers. In addition, ionization by negative‐ion chemical ionization and electrospray ionization resulted in similar spectra, which further highlights the general potential of the high‐energy CID MS/MS technique. Copyright © 2011 John Wiley & Sons, Ltd.     

Lipophilic constituents from two processed products and three different parts of Changium smyrnioides Wolff.

This article reports the lipophilic chemical composition of different processed products (Changii Radix, Changii Radix Alba) and parts (root bark, leaf and fruit) of Changium smyrnioides Wolff.. The lipophilic constituents were extracted with petroleum ether in Soxhlet apparatus, subsequently identified and determined by gas chromatography–mass spectroscopy (GC–MS). Yield of lipophilic constituents from Changii Radix (3.65%) was about three times more than Changii Radix Alba's (1.07%), which indicated processing by boiling in water had an impact on the content of lipophilic constituents. Moreover, the major compounds in different processed products and parts were found to be fatty acids and sesquiterpenes. The results are a contribution for the lipophilic chemical composition and can serve as a reference for product development of Changium smyrnioides Wolff..     

Using sheath‐liquid reagents for capillary electrophoresis‐mass spectrometry: Application to the analysis of phenolic plant extracts

The combination of CE and MS is now a widely used tool that can provide a combination of high resolution separations with detailed structural information. Recently, we highlighted the benefits of an approach to add further functionality to this well‐established hyphenated technique, namely the possibility to perform chemical reactions within the sheath‐liquid of the CE‐MS interface 1 . Apart from using hydrogen/deuterium exchange for online determination of numbers of exchangeable protons, the addition of DPPH? (2,2‐diphenyl‐1‐picrylhydrazyl) to the sheath‐liquid can be used as a fast screening tool for studying antioxidant characteristics of individual components. Such a CE‐MS methodology allows rapid and information‐rich analysis with minimal reagent and sample consumption to be performed. In the present work, we demonstrate the applicability of this approach for the characterization of phenolic plant extracts from the Labiatae family, namely Rosmarinus officinalis and Melissa officinalis. Using the described approach, a wide range of compounds (15 and 13 phenolic compounds, respectively) could be confidently identified using a combination of high resolution CE‐MS separations with implementation of online deuterium exchange and DPPH? reactions. These compounds included polyphenols, phenolic acids, and triterpene acids. In conjunction with online MS/MS experiments, extensive structural information for aglyconic and glycosylated antioxidants present in the extracts could be obtained using simple experimental changes, which can be carried out prior to the purchasing of expensive chemical standards or the time‐consuming preparative isolation of individual compounds.     

Simultaneous determination of ten flavonoids of crude and wine‐processed Radix Scutellariae aqueous extracts in rat plasma by UPLC‐ESI‐MS/MS and its application to a comparative pharmacokinetic study

Radix Scutellariae (RS) is a herbal medicine with various pharmacological activities to treat inflammation, respiratory and gastrointestinal infections, etc. In this study, a rapid, sensitive and selective UPLC‐ESI‐MS/MS method was developed for simultaneous determination of 10 flavonoids – scutellarin, scutellarein, chrysin, wogonin, baicalein, apigenin, wogonoside, oroxylin A‐7‐O‐glucuronide, oroxylin A and baicalin – from RS aqueous extracts in rat plasma with propyl paraben as internal standard (IS). Chromatographic separation was achieved on a C₁₈ column using gradient elution with the mobile phase consisting of methanol and water (containing 0.1% formic acid) at a flow rate of 0.2 mL/min. The detection was performed in multiple reaction monitoring mode using electrospray ionization in negative mode. The validated method showed good linearity over a wide concentration range (r >0.9935). The intra‐ and interday assay variabilities were <9.5% and <12.4% for all analytes, respectively. The extraction recovery ranged from 71.2 to 89.7% for each analyte and IS. This method was successfully applied to pharmacokinetic comparision after oral administration of crude and wine‐processed RS aqueous extracts. There were significant differences in some pharmacokinetic parameters of most analytes between crude and wine‐processed RS. This suggested that wine‐processing exerted effects absorption of most flavonoids. Copyright © 2014 John Wiley & Sons, Ltd.     

Characterization of chemical constituents in Zhi–Zi–Da–Huang decoction by ultra high performance liquid chromatography coupled with quadrupole time‐of‐flight mass spectrometry

A sensitive and reliable ultra high performance liquid chromatography coupled with quadrupole time‐of‐flight mass spectrometry method was established to separate and identify the chemical constituents of Zhi–Zi–Da–Huang decoction, a classic traditional Chinese medicine formula. The chromatographic separation was achieved on a Shim‐pack XR‐ODS C₁₈ column (75  × 3.0 mm, 2.2 μm) using a gradient elution program. The detection was performed on a Waters Xevo G2 Q‐TOF mass spectrometer equipped with electrospray ionization source in both positive and negative modes. With the optimized conditions, a total of 82 compounds were identified or tentatively characterized. Of the 82 compounds, 21 compounds were identified by comparing the retention time and MS data with reference standards, the rest were characterized by analyzing MS data and retrieving the reference literature. In addition, 31 compounds were identified from Gardenia jasminoides Ellis, ten compounds were identified from Rheum palmatum L., 33 compounds were identified from Citrus aurantium L., and eight compounds were identified from Sojae Semen Praeparatum. Results indicated that iridoids, anthraquinones, flavonoids, isoflavonoids, coumarins, glycosides of crocetin, monoterpenoids, and organic acids were major constituents in Zhi–Zi–Da–Huang decoction. It is concluded that the developed ultra high performance liquid chromatography coupled with quadrupole time‐of‐flight mass spectrometry method with high sensitivity and resolution is suitable for identifying and characterizing the chemical constituents of Zhi–Zi–Da–Huang decoction, and the analysis provides a helpful chemical basis for further research on Zhi–Zi–Da–Huang decoction.     

GC–MS combined with chemometric techniques for the quality control and original discrimination of Curcumae longae rhizome: Analysis of essential oils

Curcumae longae rhizome is a widely used traditional herb in many countries. Various geographical origins of this herb might lead to diversity or instability of the herbal quality. The objective of this work was to establish the chemical fingerprints for quality control and find the chemical markers for discriminating these herbs from different origins. First, chemical fingerprints of essential oil of 24 C. longae rhizome from four different geographical origins in China were determined by GC–MS. Then, pattern recognition techniques were introduced to analyze these abundant chemical data in depth; hierarchical cluster analysis was used to sort samples into groups by measuring their similarities, and principal component analysis and partial least‐squares discriminate analysis were applied to find the main chemical markers for discriminating these samples. Curcumae longae rhizome from Guangxi province had the highest essential oil yield (4.32 ± 1.45%). A total of 46 volatile compounds were identified in total. Consistent results were obtained to show that C. longae rhizome samples could be successfully grouped according to their origins, and turmerone, ar‐turmerone, and zingiberene were the characteristic components for discriminating these samples of various geographical origins and for quality control. This finding revealed that fingerprinting analysis based on GC–MS coupled with chemometric techniques could provide a reliable platform to discriminate herbs from different origins, which is a benefit for quality control.     

Rapid quality assessment of Radix Aconiti Preparata using direct analysis in real time mass spectrometry

This study presents a novel and rapid method to identify chemical markers for the quality control of Radix Aconiti Preparata, a world widely used traditional herbal medicine. In the method, the samples with a fast extraction procedure were analyzed using direct analysis in real time mass spectrometry (DART MS) combined with multivariate data analysis. At present, the quality assessment approach of Radix Aconiti Preparata was based on the two processing methods recorded in Chinese Pharmacopoeia for the purpose of reducing the toxicity of Radix Aconiti and ensuring its clinical therapeutic efficacy. In order to ensure the safety and effectivity in clinical use, the processing degree of Radix Aconiti should be well controlled and assessed. In the paper, hierarchical cluster analysis and principal component analysis were performed to evaluate the DART MS data of Radix Aconiti Preparata samples in different processing times. The results showed that the well processed Radix Aconiti Preparata, unqualified processed and the raw Radix Aconiti could be clustered reasonably corresponding to their constituents. The loading plot shows that the main chemical markers having the most influence on the discrimination amongst the qualified and unqualified samples were mainly some monoester diterpenoid aconitines and diester diterpenoid aconitines, i.e. benzoylmesaconine, hypaconitine, mesaconitine, neoline, benzoylhypaconine, benzoylaconine, fuziline, aconitine and 10-OH-mesaconitine. The established DART MS approach in combination with multivariate data analysis provides a very flexible and reliable method for quality assessment of toxic herbal medicine.     

Strategy for rapid screening of antioxidant and anti‐inflammatory active ingredients in Gynura procumbens (Lour.) Merr. based on UHPLC–Q‐TOF–MS/MS and characteristic ion filtration

Gynura procumbens (Lour.) Merr. is traditionally used as a raw material for making dumplings or steamed stuffed buns, and its fresh leaves are boiled with water for tea. Herein, we established an ultra‐high–performance liquid chromatography–quadrupole time‐of‐flight mass spectrometry (UHPLC–Q‐TOF–MS/MS) combined with characteristic ion filtration (CIF) strategy to rapidly screen active ingredients with antioxidant and anti‐inflammatory properties in G. procumbens. This strategy involved screening the active part of G. procumbens using antioxidation and anti‐inflammatory activity assays; discovering the active compounds by speculating on the active site's chemical composition by UHPLC–Q‐TOF–MS/MS plus CIF; and verifying the active compounds' activities. The ethyl acetate extract (EEAF) of G. procumbens was the major active site. Eighty‐one compounds were identified from the EEAF using UHPLC–Q‐TOF–MS/MS plus CIF. Furthermore, polyphenols such as cynarine, isochlorogenic acids A and isochlorogenic acids C have excellent antioxidizing and anti‐inflammatory activities. This study provides a practical strategy for rapid in vitro screening of the antioxidizing and anti‐inflammatory activities of traditional vegetables and herbs and identification of active ingredients.