A Support Vector Machine-Assisted Metabolomics Approach for Non-Targeted Screening of Multi-Class Pesticides and Veterinary Drugs in Maize.

The contamination risks of plant-derived foods due to the co-existence of pesticides and veterinary drugs (P&VDs) have not been fully understood. With an increasing number of unexpected P&VDs illegally added to foods, it is essential to develop a non-targeted screening method for P&VDs for their comprehensive risk assessment. In this study, a modified support vector machine (SVM)-assisted metabolomics approach by screening eligible variables to represent marker compounds of 124 multi-class P&VDs in maize was developed based on the results of high-performance liquid chromatography-tandem mass spectrometry. Principal component analysis and orthogonal partial least squares discriminant analysis indicate the existence of variables with obvious inter-group differences, which were further investigated by S-plot plots, permutation tests, and variable importance in projection to obtain eligible variables. Meanwhile, SVM recursive feature elimination under the radial basis function was employed to obtain the weight-squared values of all the variables ranging from large to small for the screening of eligible variables as well. Pairwise t-tests and fold changes of concentration were further employed to confirm these eligible variables to represent marker compounds. The results indicate that 120 out of 124 P&VDs can be identified by the SVM-assisted metabolomics method, while only 109 P&VDs can be found by the metabolomics method alone, implying that SVM can promote the screening accuracy of the metabolomics method. In addition, the method's practicability was validated by the real contaminated maize samples, which provide a bright application prospect in non-targeted screening of contaminants. The limits of detection for 120 P&VDs in maize samples were calculated to be 0.3~1.5 µg/kg.

Development and Validation of a High-Performance Liquid Chromatography Method to Quantify Marker Compounds in Lysimachia vulgaris var. davurica and Its Effects in Diarrhea-Predominant Irritable Bowel Syndrome.

Irritable bowel syndrome (IBS), a common gastrointestinal disorder worldwide, is characterized by chronic abdominal pain, bloating, and disordered defecation. IBS is associated with several factors, including visceral hypersensitivity, gut motility, and gut-brain interaction disorders. Because currently available pharmacological treatments cannot adequately improve symptoms and may cause adverse effects, the use of herbal therapies for managing IBS is increasing. Lysimachia vulgaris var. davurica (LV) is a medicinal plant used in traditional medicine to treat diarrhea. However, information on whether LV can effectively improve diarrhea-predominant IBS (IBS-D) remains limited. In this study, using an experimental mouse model of IBS-D, we elucidated the effects of the LV extract. The methanol extract of LV decreased fecal pellet output in the restraint stress- or 5-hydroxytryptamine (5-HT)-induced IBS mouse model and inhibited 5-HT-mediated [Ca2+]i increase in a dose-dependent manner. Furthermore, we developed and validated a high-performance liquid chromatography method using two marker compounds, namely, chlorogenic acid and rutin, for quality control analysis. Our study results suggest the feasibility of the methanol extract of LV for developing therapeutic agents to treat IBS-D by acting as a 5-HT3 receptor antagonist.

Identification of antioxidant and flavour marker compounds in Kalosi-Enrekang Arabica brewed coffee processed using different postharvest treatment methods.

This research aims to predict the presence of marker compounds that differentiate tubruk brew from coffee beans with different postharvest processing. This research also aims to predict compounds correlating with antioxidant activity and sensory flavour attributes. This research used Kalosi-Enrekang Arabica coffee beans, which were processed with three different postharvest processing (honey, full-washed and natural), roasted at medium level, and brewed using the tubruk method. Each brew was analyzed for chemical profiles using LC-MS and GC-MS, antioxidant analysis using the DPPH IC50 and FRAP methods, and sensory analysis for flavour using the QDA and SCAA methods for cupping scores. OPLS-DA analysis revealed the presence of marker compounds from each brew, and the dried fruit flavour attribute was to be an inter-process marker. After that, OPLS analysis showed marker compounds that correlate to antioxidant activity and flavour attributes. Rhaponticin is thought to be one of the marker compounds in natural coffee brews and is one of the compounds that correlates to the antioxidant activity of the DPPH method (IC50); prunin is thought to be one of the marker compounds for full-washed coffee brews and is one of the compounds that correlates to the activity antioxidants of FRAP method. Triacetin, which is thought to be a marker compound in natural brewed coffee, correlates with fruity flavour. 3-acetylpyridine, as a marker in honey-brewed coffee, correlates with nutty flavour. Even though there are differences in dominant flavours, the cupping score shows the brew is categorized as a specialty. This research shows that different post-harvest processing processes influence the compound profile, antioxidant activity and flavour attributes of Tubruk brewed coffee. Supplementary Information: The online version contains supplementary material available at 10.1007/s13197-024-05948-8.

Quality Assessment and Classification of Codonopsis Radix Based on Fingerprints and Chemometrics.

In China, Codonopsis Radix (CR) is frequently consumed both as food and medicine. Here, a comprehensive strategy based on fingerprinting and chemometric approaches was created to explore the influence of origins, storage time and kneading processing on the quality of CR. Firstly, high-performance liquid chromatography with diode array detection was used to obtain the fingerprints of 35 batches of CR from six different origins and 33 batches of CR from varying storage times or kneading procedures. Secondly, chemometric methods including similarity analysis (SA), principal component analysis (PCA), hierarchical clustering analysis (HCA), and two-way orthogonal partial least square with discriminant analysis (O2PLS-DA) were used to evaluate the differences of chemical components in CR so as to identify its source and reflect its quality. Moreover, 13 and 16 major compounds were identified as marker compounds for the discrimination of CR from different origins, storage time and kneading processing, respectively. Furthermore, the relative content of the marker components and the exact content of Lobetyolin were measured, indicating that the contents of these components vary significantly between various CR samples. Meanwhile, the chemical components of CR were identified using Mass spectrometry. According to the findings of our investigation, the quality of CR from Gansu was the best, followed by Shanxi and then Sichuan. The quality of CR from Chongqing and Guizhou was poor. At the same time, the quality of CR was the best when it was kneaded and stored for 0 years, indicating that the traditional kneading process of CR is of great significance. Conclusively, HPLC fingerprint in conjunction with chemical pattern recognition and component content determination can be employed to differentiate the raw materials of different CR samples. Additionally, it is also a reliable, comprehensive and prospective method for quality control and evaluation of CR.

The Efficient Synthesis and Anti-Fatigue Activity Evaluation of Macamides: The Unique Bioactive Compounds in Maca.

Macamides are a class of amide alkaloids that are only found in maca and are widely considered to be its bioactive marker compounds. More than thirty macamide monomers have been identified in recent years; however, it is difficult to obtain a single macamide monomer from the maca plant because of their similar structures and characteristics. We used the carbodiimide condensation method (CCM) to efficiently synthesize five typical macamides, including N-benzyl-hexadecanamide (NBH), N-benzyl-9Z,12Z,15Z-octadecenamide, N-(3-methoxybenzyl)-9Z,12Z-octadecenamide, N-benzyl-9Z,12Z-octadecenamide, and N-(3-methoxybenzyl)-9Z,12Z,15Z-octadecadienamide. All the synthesized macamides were purified by a one-step HPLC with a purity of more than 95%. NBH is the most abundant macamide monomer in natural maca, and it was selected to evaluate the anti-fatigue effects of macamides. The results indicated that NBH could enhance the endurance capacity of mice by increasing liver glycogen levels and decreasing blood urea nitrogen, lactate dehydrogenase, blood ammonia, and blood lactic acid levels. Macamides might be the active substances that give maca its anti-fatigue active function.

Identification of Daphne genkwa and Its Vinegar-Processed Products by Ultraperformance Liquid Chromatography-Quadrupole Time-of-Flight Mass Spectrometry and Chemometrics.

Crude herbs of Daphne genkwa (CHDG) are often used in traditional Chinese medicine to treat scabies baldness, carbuncles, and chilblain owing to their significant purgation and curative effects. The most common technique for processing DG involves the use of vinegar to reduce the toxicity of CHDG and enhance its clinical efficacy. Vinegar-processed DG (VPDG) is used as an internal medicine to treat chest and abdominal water accumulation, phlegm accumulation, asthma, and constipation, among other diseases. In this study, the changes in the chemical composition of CHDG after vinegar processing and the inner components of the changed curative effects were elucidated using optimized ultrahigh-performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry (UPLC-Q-TOF-MS). Untargeted metabolomics, based on multivariate statistical analyses, was also used to profile differences between CHDG and VPDG. Eight marker compounds were identified using orthogonal partial least-squares discrimination analysis, which indicated significant differences between CHDG and VPDG. The concentrations of apigenin-7-O-ß-d-methylglucuronate and hydroxygenkwanin were considerably higher in VPDG than those in CHDG, whereas the amounts of caffeic acid, quercetin, tiliroside, naringenin, genkwanines O, and orthobenzoate 2 were significantly lower. The obtained results can indicate the transformation mechanisms of certain changed compounds. To the best of our knowledge, this study is the first to employ mass spectrometry to detect the marker components of CHDG and VPDG.

Propolis: chemical diversity and challenges in quality control.

Propolis is a resinous natural product produced by honeybees using beeswax and plant exudates. The chemical composition of propolis is highly complex, and varies with region and season. This inherent chemical variability presents several challenges to its standardisation and quality control. The present review was aimed at highlighting marker compounds for different types of propolis, produced by the species Apis mellifera, from different geographical origins and that display different biological activities, and to discuss strategies for quality control. Over 800 compounds have been reported in the different propolises such as temperate, tropical, birch, Mediterranean, and Pacific propolis; these mainly include alcohols, acids and their esters, benzofuranes, benzopyranes, chalcones, flavonoids and their esters, glycosides (flavonoid and diterpene), glycerol and its esters, lignans, phenylpropanoids, steroids, terpenes and terpenoids. Among these, flavonoids (> 140), terpenes and terpenoids (> 160) were major components. A broad range of biological activities, such as anti-oxidant, antimicrobial, anti-inflammatory, immunomodulatory, and anticancer activities, have been ascribed to propolis constituents, as well as the potential of these compounds to be biomarkers. Several analytical techniques, including non-separation and separation methods have been described in the literature for the quality control assessment of propolis. Mass spectrometry coupled with separation methods, followed by chemometric analysis of the data, was found to be a valuable tool for the profiling and classification of propolis samples, including (bio)marker identification. Due to the rampant chemotypic variability, a multiple-marker assessment strategy considering geographical and biological activity marker(s) with chemometric analysis may be a promising approach for propolis quality assessment. Supplementary Information: The online version contains supplementary material available at 10.1007/s11101-022-09816-1.

Metabolomics Approach on Non-Targeted Screening of 50 PPCPs in Lettuce and Maize.

The metabolomics approach has proved to be promising in achieving non-targeted screening for those unknown and unexpected (U&U) contaminants in foods, but data analysis is often the bottleneck of the approach. In this study, a novel metabolomics analytical method via seeking marker compounds in 50 pharmaceutical and personal care products (PPCPs) as U&U contaminants spiked into lettuce and maize matrices was developed, based on ultrahigh-performance liquid chromatography-tandem mass spectrometer (UHPLC-MS/MS) output results. Three concentration groups (20, 50 and 100 ng mL-1) to simulate the control and experimental groups applied in the traditional metabolomics analysis were designed to discover marker compounds, for which multivariate and univariate analysis were adopted. In multivariate analysis, each concentration group showed obvious separation from other two groups in principal component analysis (PCA) and orthogonal partial least squares discriminant analysis (OPLS-DA) plots, providing the possibility to discern marker compounds among groups. Parameters including S-plot, permutation test and variable importance in projection (VIP) in OPLS-DA were used for screening and identification of marker compounds, which further underwent pairwise t-test and fold change judgement for univariate analysis. The results indicate that marker compounds on behalf of 50 PPCPs were all discovered in two plant matrices, proving the excellent practicability of the metabolomics approach on non-targeted screening of various U&U PPCPs in plant-derived foods. The limits of detection (LODs) for 50 PPCPs were calculated to be 0.4~2.0 µg kg-1 and 0.3~2.1 µg kg-1 in lettuce and maize matrices, respectively.

HPLC-UV/HRMS methods for the unambiguous detection of adulterations of Ginkgo biloba leaves with Sophora japonica fruits on an extract level.

CONTEXT: Ginkgo biloba L. (Ginkgoaceae) leaf extract is one of the most frequently sold herbal extracts. There have been reports on poor quality and adulteration of ginkgo leaf extracts or the powdered plant material with extracts or powder of Styphnolobium japonicum (L.) Schott (Fabaceae) (syn. Sophora japonica L.) fruits, which is rich in flavone glycosides. OBJECTIVE: The study investigates whether ginkgo leaves genuinely contain genistein and sophoricoside and whether these two substances could be used as markers to detect adulterations with sophora fruits. MATERIALS AND METHODS: A total of 33 samples of dried ginkgo leaves were sourced from controlled plantations in China, the USA, and France. After extraction, the samples were analyzed using two high-performance liquid chromatography (HPLC) coupled with UV/HRMS methods for the detection of genistein and sophoricoside, respectively. Chromatograms were compared to standard reference materials. RESULTS: In none of the tested ginkgo samples, neither genistein nor sophoricoside could be detected. The applied method was designed to separate genistein from apigenin. The latter is a genuine compound of ginkgo leaves, and its peak may have been previously misidentified as genistein because of the same molecular mass. The method for the detection of sophoricoside allows identification of the adulteration with sophora fruit without prior hydrolysis. By both HPLC methods, it was possible to detect adulterations of ≥2% sophora fruits in the investigated ginkgo extract. CONCLUSION: The methods allow unambiguous detection of adulterations of ginkgo leaves with sophora fruits, using genistein and sophoricoside as marker compounds.

Honey Volatiles as a Fingerprint for Botanical Origin-A Review on their Occurrence on Monofloral Honeys.

Honeys have specific organoleptic characteristics, with nutritional and health benefits, being highly appreciated by consumers, not only in food but also in the pharmaceutical and cosmetic industries. Honey composition varies between regions according to the surrounding flora, enabling its characterization by source or type. Monofloral honeys may reach higher market values than multifloral ones. Honey's aroma is very specific, resulting from the combination of volatile compounds present in low concentrations. The authentication of honey's complex matrix, according to its botanical and/or geographical origin, represents a challenge nowadays, due to the different sorts of adulteration that may occur, leading to the search for reliable marker compounds for the different monofloral honeys. The existing information on the volatiles of monofloral honeys is scarce and disperse. In this review, twenty monofloral honeys and honeydews, from acacia, buckwheat, chestnut, clover, cotton, dandelion, eucalyptus, fir tree, heather, lavender, lime tree, orange, pine, rape, raspberry, rhododendron, rosemary, strawberry tree, sunflower and thyme, were selected for volatile comparison purposes. Taking into consideration the country of origin, the technique of isolation and analysis, the five main volatiles from each of the honeys are compared. Whereas some compounds were found in several types of monofloral honey, and thus not considered good volatile markers, some monofloral honeys revealed characteristic volatile compounds independently of their provenance.

Similarity and differential NMR spectroscopy in metabolomics: application to the analysis of vegetable oils with 1H and 13C NMR.

INTRODUCTION: In NMR based metabolomics there is a need for tools to easily compare spectra and to extract the maximum of information from the data. OBJECTIVES: The calculation of similarity and performing differential NMR spectroscopy provides important additional information for classification and validation in metabolomics experiments. METHODS: From 13 different vegetable oils samples were analysed by 1H and 13C NMR. The similarity between spectra was calculated and differential NMR spectroscopy was used to discover marker compounds. RESULTS: The similarity between the individual spectra was calculated for the spectra of all samples. The similarity was used to verify and improve the alignment. For vegetable oils which showed a high similarity, e.g. chia seed oil and linseed oil, differential NMR spectroscopy was used to discover marker compounds. CONCLUSIONS: The calculation of similarity is an important tool to reveal variability between samples and spectra and can be used to verify data sets and improve alignment or binning procedures. With differential spectroscopy marker compounds are easily discovered. The methods can be seen as an important addition to the routine procedures of metabolomics experiments.

Comparison of Phytochemical Differences of the Pulp of Different Peach [Prunus persica (L.) Batsch] Cultivars with Alpha-Glucosidase Inhibitory Activity Variations in China Using UPLC-Q-TOF/MS.

In order to fully understand the variation of the fruit alpha-glucosidase inhibitory activity-related phytochemical basis in the Chinese peach [Prunus persica (L.) Batsch], mature fruit from 33 cultivars was used for the investigation of fruit phenolic phytochemical attributes, including total phenolics, flavonoids, anthocyanins, and procyanidins, as well as the alpha-glucosidase inhibitory activity in vitro. Alpha-glucosidase inhibitory activity varied significantly among tested peach cultivars and was strongly correlated with total phenolics, total procyanidins, and total flavonoids. Untargeted UPLC-Q-TOF/MS-based metabolomics were used to comprehensively discriminate between peaches with different inhibitory activity on alpha-glucosidase. Principal component analysis (PCA) and orthogonal partial least squares discrimination analysis (OPLS-DA) were used for this process. Twenty-three differential compounds were identified between peach cultivars with high and low alpha-glucosidase inhibitory activity, and nine, including procyanidin C1, procyanidin trimer isomer 1, procyanidin trimer isomer 2, procyanidin B1, procyanidin dimer, epicatechin-epicatechin-epicatechin, phloridzin, kaempferol 3-(2'',6''-di-(E)-p-coumarylglucoside), and luteolin 3'-methyl ether 7-malonylglucoside, were identified as marker compounds responsible for the discrimination. Overall, variations in metabolites in peach pulp reflect the diversity in peach germplasm, and these nine compounds are good candidate markers for future genetic breeding of peach fruit with high alpha-glucosidase inhibitory activity.

Inferring the origin of rare fruit distillates from compositional data using multivariate statistical analyses and the identification of new flavour constituents.

BACKGROUND: In Serbia, delicatessen fruit alcoholic drinks are produced from autochthonous fruit-bearing species such as cornelian cherry, blackberry, elderberry, wild strawberry, European wild apple, European blueberry and blackthorn fruits. There are no chemical data on many of these and herein we analysed volatile minor constituents of these rare fruit distillates. Our second goal was to determine possible chemical markers of these distillates through a statistical/multivariate treatment of the herein obtained and previously reported data. RESULTS: Detailed chemical analyses revealed a complex volatile profile of all studied fruit distillates with 371 identified compounds. A number of constituents were recognised as marker compounds for a particular distillate. Moreover, 33 of them represent newly detected flavour constituents in alcoholic beverages or, in general, in foodstuffs. With the aid of multivariate analyses, these volatile profiles were successfully exploited to infer the origin of raw materials used in the production of these spirits. It was also shown that all fruit distillates possessed weak antimicrobial properties. CONCLUSION: It seems that the aroma of these highly esteemed wild-fruit spirits depends on the subtle balance of various minor volatile compounds, whereby some of them are specific to a certain type of fruit distillate and enable their mutual distinction.

Chemical interaction between Paeonia lactiflora and Glycyrrhiza uralensis, the components of Jakyakgamcho-tang, using a validated high-performance liquid chromatography method: herbal combination and chemical interaction in a decoction.

The herbal combination is the basic unit of a herbal formula that affects the chemical characteristics of individual herbs. In the present study, a method of simultaneous determination of the 11 marker compounds in Jakyakgamcho-tang was developed using high-performance liquid chromatography with photodiode array detection. The validated analytical method was successfully applied to approach the chemical interaction between Paeonia lactiflora and Glycyrrhiza uralensis in co-decoction. In P. lactiflora, the contents of gallic acid, oxypaeoniflorin, (+)-catechin, paeoniflorin, and benzoylpaeoniflorin were decreased, while those of albiflorin and benzoic acid were increased; in G. uralensis, the contents of liquiritin, isoliquiritin, ononin, and glycyrrhizin were decreased, when decocting two herbs together. Moreover, as the ratio between P. lactiflora and G. uralensis was increased, the contents of chemical contents from each herb were proportionally increased. However, each content of marker compound per the gram of herbal medicine was decreased as the ratio of combinative herbs increased. The results showed that P. lactiflora and G. uralensis affect the extraction efficiency of chemical compounds in a Jakyakgamcho-tang decoction. Overall, the method established in this study was simple, rapid, and accurate, and would be useful for the determination of marker compounds and for the investigation of the chemical interaction between herbal medicines.

Application of HS-SPME-GC-MS combined with electronic nose technology in the odor recognition of pseudostellariae radix.

Correct identification of the substance basis of Pseudostellariae Radix (PR) odor is important not only for the quality control of the products, but also for the safety of the consumers. PR is often described with a special smell, such as strange, moldy or earthy. Electronic nose-based technology coupled with headspace solid phase microextraction gas chromatography-mass spectrometry (HS-SPME-GC-MS) was used to investigate the volatile components in PR from 47 germplasms cultivated in traditional fields. A total of 48 common compounds were identified based on HS- SPME-GC-MS technology, and 25 of them with aroma characteristics were found based on Alpha soft 13.4. The 1-Octen-3-ol, geosim, (E)-2-nonenal and 1- methylnaphthalene as contributing marker compounds of the 'specific smell' of PR were identified. The odor recognition mode, with demonstrated excellent accuracy in recognition abilities, enabled the correct identification of commercial samples including complex mixtures.

Insight from untargeted metabolomics: Revealing the potential marker compounds changes in refrigerated pork based on random forests machine learning algorithm.

Data on changes in non-volatile components and metabolic pathways during pork storage were inadequately investigated. Herein, an untargeted metabolomics coupled with random forests machine learning algorithm was proposed to identify the potential marker compounds and their effects on non-volatile production during pork storage by ultra-high-performance liquid chromatography-mass spectrometry (UHPLC-MS/MS). A total of 873 differential metabolites were identified based on analysis of variance (ANOVA). Bioinformatics analysis shows that the key metabolic pathways for protein degradation and amino acid transport are amino acid metabolism and nucleotide metabolism. Finally, 40 potential marker compounds were screened using the random forest regression model, innovatively proposing the key role of pentose-related metabolism in pork spoilage. Multiple linear regression analysis revealed that d-xylose, xanthine, and pyruvaldehyde could be key marker compounds related to the freshness of refrigerated pork. Therefore, this study could provide new ideas for the identification of marker compounds in refrigerated pork.

Multivariate Analysis among Marker Compounds, Environmental Factors, and Fruit Quality of Schisandra chinensis at Different Locations in South Korea.

This study aimed to investigate the correlation among the contents of marker compounds, growth characteristics, and environmental factors of Schisandra chinensis fruits across South Korea. The fruits were collected from 36 cultivation sites in 28 regions across the country. We investigated nine growth characteristics, twelve soil physicochemical properties, eight meteorological data, and three marker compounds in this study. We optimized and validated an optimized method for quantifying marker compounds using UPLC and performed correlation analysis among the contents of marker compounds, growth characteristics, and environmental factors. The UPLC-UV method for analyzing marker compounds was validated by measuring linearity, LOD, LOQ, precision, and accuracy. The marker compounds were negatively correlated with the fruit size and sugar contents, and growth characteristics were negatively correlated with some physicochemical properties of the soil. The results of this study can be used as basic data for the standard cultural practices and quality control of S. chinensis fruits.

Licochalcone L, an undescribed retrochalcone from Glycyrrhiza inflata roots.

Glycyrrhiza inflata Batalin is among the three glycyrrhizin producing Glycyrrhiza species and can be distinguished from other species with regard to its retrochalcone contents. Seven retrochalcones, echinatin and licochalcones A, C, D, E, K, and L were isolated and characterized from the chloroform extract of G. inflata roots. Among the isolates, licochalcone L was found to be previously undescribed. Structure elucidation of these specialised metabolites was achieved through NMR and mass spectroscopic data analyses.

Geographical differentiation of Molixiang table grapes grown in China based on volatile compounds analysis by HS-GC-IMS coupled with PCA and sensory evaluation of the grapes.

In this study, the volatile fingerprints of GC-IMS showed great differences on the volatile profiles of Molixiang grapes collected from three different regions of China, which suggested that aroma of table grapes could be largely impacted by origin areas. Butyl lactate, E-2-octenal and Z-2-pentanol were mainly contained in MLX-A, the grapes sampled from Ningbo, China. High contents of p-cymene, styrene and γ-terpinene were observed in MLX-B grapes sampled from Beizhen, China. In addition, benzaldehyde and methyl benzoate were major contained in MLX-C grapes sampled from Zhangzhou, China. The PCA results revealed effective differentiation of samples from different geographical origin based on the information obtained from GC-IMS. Furthermore, sensory evaluation showed that the aroma characters of grapes from different geographical origin were significantly different (P ≤ 0.05). E-2-octenal, styrene and benzaldehyde might serve as the geographical marker compounds of origin area based on the results of GC-IMS analysis and sensory evaluation.