Comparison of Approaches for Authentication of Commercial Terpinen-4-ol-type Tea Tree Oils Using Chiral GC/MS.

A global demand for tea tree oil (TTO) has resulted in increased adulteration in commercial products. In this study, we use a novel enantiomeric gas chromatography mass spectrometry method for chiral analysis of key terpenes ((±)-terpinen-4-ol, (±)-α-terpineol, and (±)-limonene) and quantification of components present at >0.01% to test different methods of identifying adulterated TTO. Data from authentic Australian (n = 88) and oxidized (n = 12) TTO samples of known provenance were consistent with recommended ranges in ISO 4730:2017 and previously published enantiomeric ratios, with p-cymene identified as the major marker of TTO oxidation. The 15 ISO 4730:2017 constituents comprised between 84.5 and 89.8% of the total ion chromatogram (TIC) peak area. An additional 53 peaks were detected in all samples (7.3-11.0% of TIC peak area), while an additional 43 peaks were detected in between 0 and 99% (0.15-2.0% of the TIC peak area). Analysis of nine commercial samples demonstrated that comparison to the ISO 4730:2017 standard does not always identify adulterated TTO samples. While statistical analysis of minor components in TTO did identify two commercial samples that differed from authentic TTO, the (+)-enantiomer percentages for limonene, terpinen-4-ol, and α-terpineol provided clearer evidence that these samples were adulterated. Thus, straightforward identification of unadulterated and unoxidized TTO could be based on analysis of appropriate enantiomeric ratios and quantitation of the p-cymene percentage.

Intelligent chemical profiling of 73 edible flowers by liquid chromatography-high resolution mass spectrometry combined with HRMS database and their authentication based on large-scale fingerprints.

A commercial high-resolution MS database "TCM-PCDL" was innovatively introduced to automatically identify multi-components in 73 edible flowers rapidly and accurately by liquid chromatography-high resolution mass spectrometry, which can be time-consuming and labor-intensive in traditional manual method. The database encompasses over 2565 natural products with various energy levels. Unknown compounds can be identified through direct matching and scoring MS2 spectra with database. A total of 870 compounds were identified from 73 flowers, with polyphenols constituting up to 75%. Focusing on polyphenols, a high performance liquid chromatography (HPLC) method was developed to generate fingerprints from 510 batches, establishing an "HPLC database" that enabled accurate authentication using similarity scores and rankings. This method demonstrated an accuracy rate of 100% when applied to 30 unknown samples. For flowers prone to confusion, additional statistical analysis methods could be employed as aids in authentication. This study provides valuable insights for large-scale sample chemical profiling and authentication.

Machine learning-enabled high-throughput industry screening of edible oils.

Long-term consumption of mixed fraudulent edible oils increases the risk of developing of chronic diseases which has been a threat to the public health globally. The complicated global supply-chain is making the industry malpractices had often gone undetected. In order to restore the confidence of consumers, traceability (and accountability) of every level in the supply chain is vital. In this work, we shown that machine learning (ML) assisted windowed spectroscopy (e.g., visible-band, infra-red band) produces high-throughput, non-destructive, and label-free authentication of edible oils (e.g., olive oils, sunflower oils), offers the feasibility for rapid analysis of large-scale industrial screening. We report achieving high-level of discriminant (AUC > 0.96) in the large-scale (n ≈ 11,500) of adulteration in olive oils. Notably, high clustering fidelity of 'spectral fingerprints' achieved created opportunity for (hypothesis-free) self-sustaining large database compilation which was never possible without machine learning. (137 words).

Three-dimensional fluorescence combined with alternating trilinear decomposition and random forest algorithm for the rapid prediction of species, geographical origin and main components of Glycyrrhizae Radix et Rhizoma (Gancao).

Glycyrrhizae Radix et Rhizoma (Gancao) is a functional food whose quality varies significantly between distinct geographical sources owing to the influence of genetics and the geographical environment. This study employed three-dimensional fluorescence coupled with alternating trilinear decomposition (ATLD) and random forest (RF) algorithms to rapidly predict Gancao species, geographical origins, and primary constituents. Seven fluorescent components were resolved from the three-dimensional fluorescence of the ATLD for subsequent analysis. Results indicated that the RF model distinguished Gancao from various species and origins better than other algorithms, achieving an accuracy of 94.4 % and 88.9 %, respectively. Furthermore, the RF regressor algorithm was used to predict the concentrations of liquiritin and glycyrrhizic acid in Gancao, with 96.4 % and 95.6 % prediction accuracies compared to HPLC, respectively. This approach offers a novel means of objectively evaluating the origin of food and holds substantial promise for food quality assessment.

Identification of authentic Phyllanthus amarus species in India.

The Phyllanthus is a plant used in the traditional Ayurvedic Medicine system and has more than 800 species. These species grow in the same area and there are chances of adulteration of other species and, incorrect identification may also lead to wrong reports. This study was attempted by Enovate Biolife Pvt. Ltd. to identify authentic Phyllanthus amarus. The nine raw material samples were collected from different populations/suppliers from various zones of India for the study. All the samples were analysed using microscopic and macroscopic ID, and by using the High Performance Thin Layer Chromatography (HPTLC) fingerprint method. The samples collected from the Central zone (Lucknow PA-08, Uttar Pradesh) and the Southern zone (Coimbatore PA-05, and Chennai PA-09, Tamil Nadu) of India were found to be authentic P. amarus by the mentioned identification methods.

Integration of untargeted and pseudotargeted metabolomics reveals specific markers for authentication and adulteration detection of Fritillariae Bulbus using tandem mass spectrometry and chemometrics.

Authentication and adulteration detection of closely related herbal medicines is a thorny issue in the quality control and market standardization of traditional Chinese medicine. Taking Fritillariae Bulbus (FB) as a case study, we herein proposed a three-step strategy that integrates mass spectrometry-based metabolomics and multivariate statistical analysis to identify specific markers, thereby accurately identifying FBs and determining the adulteration level. First, an ultra-high performance liquid chromatography-quadrupole time-of-flight mass spectrometry-based untargeted metabolomics method was employed to profile steroid alkaloids in five sorts of FB and screen potential differential markers. Then, the reliability of the screened markers was further verified by the distribution in different FB groups acquired from ultra-high performance liquid chromatography triple quadrupole mass spectrometry-based pseudotargeted metabolomics analysis. As a result, a total of 16 compounds were screened out to be the specific markers, which were successfully applied to distinguish five FBs by using discriminant analysis model. Besides, partial least squares regression models based on specific markers allowed accurate prediction of three sets of adulterated FBs. All the models afforded good linearity and good predictive ability with regression coefficient of prediction (R2p) > 0.9 and root mean square error of prediction (RMSEP) < 0.1. The reliable results of discriminant and quantitative analysis revealed that this proposed strategy could be potentially used to identify specific markers, which contributes to rapid chemical discrimination and adulteration detection of herbal medicines with close genetic relationship.

Phytochemistry of Red Ginseng, a Steam-Processed Panax ginseng.

Asian ginseng, the root of Panax ginseng C.A. Meyer, occupies a prominent position in the list of best-selling natural products in the world. There are two major types of ginseng roots: white ginseng and red ginseng, each with numerous preparations. White ginseng is prepared by air-drying fresh Asian ginseng roots after harvest. Red ginseng is prepared by steaming roots in controlled conditions using fresh or raw Asian ginseng. Red ginseng is commonly used in Asian countries due to its unique chemical profile, different therapeutic efficacy, and increased stability. Compared with the widespread research on white ginseng, the study of red ginseng is relatively limited. In this paper, after a botanical feature description, the structures of different types of constituents in red ginseng are systematically described, including naturally occurring compounds and those resulting from the steam processing. In red ginseng phytochemical studies, the number of published reports on ginsenosides is significantly higher than that for other constituents. Up to now, 57 ginsenosides have been isolated and characterized in red ginseng. The structural transformation pathways during steaming have been summarized. In comparison with white ginseng, red ginseng also contains other constituents, including polyacetylenes, Maillard reaction products, other types of glycosides, lignans, amino acids, fatty acids, and polysaccharides, which have also been presented. Appropriate analytical methods are necessary for differentiating between unprocessed white ginseng and processed red ginseng. Specific marker compounds and chemical profiles have been used to discriminate red ginseng from white ginseng and adulterated commercial products. Additionally, a brief phytochemical profile comparison has been made between white ginseng and black ginseng, and the latter is another type of processed ginseng prepared from white or red ginseng by steaming several times. In conclusion, to ensure the safe and effective use of red ginseng, phytochemical and analytical studies of its constituents are necessary and even crucial.

Traceability and authentication in agri-food production: A multivariate approach to the characterization ofthe Italian food excellence elephant garlic (Allium ampeloprasum L.), a vasoactive nutraceutical.

A research platform for food authentication was set up by combining stable isotope ratio analysis, metabolomics by gas and liquid mass-spectrometry and NMR investigations, chemometric analyses for food excellences. This multi-analytical approach was tested on samples of elephant garlic (Allium ampeloprasum L.), a species belonging to the same genus of common garlic (Allium ampeloprasum L.), mainly produced in southern Tuscany-(Allium ampeloprasum). The isotopic composition allowed the product to be geographically characterized. Flavonoids, like (+)-catechin, cinnamic acids, quercetin glycosides were identified. The samples showed also a significant amount of dipeptides, sulphur-containing metabolites and glutathione, the latter of which could be considered a molecular marker of the analyzed elephant garlic. For nutraceutical profiling to reach quality labels, extracts were investigated in specific biological assays, displaying interesting vasorelaxant properties in rat aorta by mediating nitric oxide release from the endothelium and exhibited positive inotropic and negative chronotropic effects in rat perfused heart.

Fluorescence lifetime imaging microscopy as a tool to characterize spice powder variations for quality and authenticity purposes: A ginger case study.

Spices are usually ground for applications and the resulting particle size of the powders is an important product attribute in view of the release of flavour. However, inhomogeneity of the original material may lead to variations in the physicochemical characteristics of the particles. This variation and its linkage to particle size may be examined by particular imaging techniques. This study aimed to explore the potential of Fluorescence Lifetime Imaging Microscopy (FLIM) to characterize spice powders according to particle size variations and correlation with their pigment contents to reveal the chemical information contained within the FLIM data. Ginger powder was used as a representative powder model. The FLIM profiles of the individual samples and populations revealed that FLIM coupled with the phasor approach has the capacity to characterize spice powder according to particle size. Meanwhile, Principal Component Analysis of pre-processed FLIM data revealed clustering of particle size groups. Further correlation analysis between the pigment compound contents and FLIM data of the ginger powders indicated that FLIM reflected chemical information of ginger powder and was able to visualize endogenous fluorophores. The current study revealed the potential of FLIM to characterize ginger powder particles. This approach may be extrapolated to other spice powder products. The new knowledge is a step further in paving the way for the application of innovative techniques, already prevalent in other domains, to food quality and authentication.

Existing status and future advancements of adulteration detection techniques in herbal products.

BACKGROUND: Herbal products have been commonly used all over the world for centuries. Its products have gained remarkable acceptance as therapeutic agents for a variety of disorders. However, following recent research disclosing discrepancies between labeling and actual components of herbal products, there is growing concern about the efficacy, quality and safety of the products. The admixture and adulteration of herbal medicinal products pose a risk of serious health compromise and the well-being of the consumers. To prevent adulteration in raw ingredients and final herbal products, it is necessary to use approaches to assess both genomes as well as metabolomics of the products; this offers quality assurance in terms of product identification and purity. The combinations of molecular and analytical methods are inevitable for thorough verification and quality control of herbal medicine. METHODS AND RESULTS: This review discusses the combination of DNA barcoding, DNA metabarcoding, mass spectroscopy as well as HPLC for the authentication of herbal medicine and determination of the level of adulteration. It also discusses the roles of PCR and real-time PCR techniques in validating and ensuring the quality, purity and identity of the herbal products. CONCLUSIONS: In conclusion, each technique has its own pros and cons, but the cumulative of both the chemical and molecular methods is proven to be the best strategy for adulteration detection. Moreover, CRISPR diagnosis tools equipped with multiplexing techniques may be implemented for screening adulteration from herbal drugs, this will play a crucial role in herbal product authentication in the future.

Development of sequential online extraction electrospray ionization mass spectrometry for accurate authentication of highly-similar Atractylodis Macrocephalae.

The accurate and rapid authentication techniques and strategies for highly-similar foods are still lacking. Herein, a novel sequential online extraction electrospray ionization mass spectrometry (S-oEESI-MS) was developed to achieve spatio-temporally resolved ionization and comprehensive characterization of complex foods with multi-components (high, medium, and low polarity substances). Meanwhile, a characteristic marker screening method and an integrated research strategy based on MS fingerprinting, characteristic marker and chemometrics modeling were established, which are especially suitable for the accurate and rapid authentication of highly-similar foods that are difficult to be authenticated by traditional techniques (e.g., LC-MS). Thirty-two batches of highly-similar Atractylodis macrocephalae rhizome from four different origins were used as model samples. As a result, S-oEESI-MS enabled a more comprehensive MS characterization of substance profiles in complex plant samples in 1.0 min. Further, 22 characteristic markers of Atractylodis macrocephalae were ingeniously screened out and combined with multivariate statistical analysis model, the accurate authentication of highly-similar Atractylodis macrocephalae was realized. This study presents a comprehensive strategy for accurate authentication and origin analysis of highly-similar foods, which has potentially significant applications for ensuring food quality and safety.

Authentication and discrimination of new Brazilian Canephora coffees with geographical indication using a miniaturized near-infrared spectrometer.

New Brazilian Canephora coffees (Conilon and Robusta) of high added value from specific origins have been protected by geographical indication to guarantee their origin and quality. Recently, benchtop near-infrared (NIR) spectroscopy combined with chemometrics has demonstrated its usefulness to discriminate them. It was the first study, however, and therefore the possibility exists to develop a new portable NIR method for this purpose. This work assessed a miniaturized NIR as a cheaper spectrometer to discriminate and authenticate new Brazilian Canephora coffees with certified geographical origins and to differentiate them from specialty Arabica. Discriminant chemometric and class modeling techniques have been applied and have obtained good predictive ability on external test sets. In addition, models with similar classification purpose were compared with those obtained in previous research carried out with benchtop NIR for the same samples, obtaining comparable results. In this context, the portable method was used as a laboratory technique and has the advantage of being cheaper than benchtop NIR spectrometer. Furthermore, it brings a high possibility to be implemented in small coffee cooperatives, industries or control agencies in the future that do not have high economic resources.

Integration of non-targeted multicomponent profiling, targeted characteristic chromatograms and quantitative to accomplish systematic quality evaluation strategy of Huo-Xiang-Zheng-Qi oral liquid.

Huo-Xiang-Zheng-Qi oral liquid (HXZQOL) is a well-known traditional Chinese medicine formula for the treatment of gastrointestinal diseases, with the pharmacologic effects of antiinflammatory, immune protection and gastrointestinal motility regulation. More significantly, HXZQOL is recommended for the treatment of COVID-19 patients with gastrointestinal symptoms, and it has been clinically proven to reduce the inflammatory response in patients with COVID-19. However, the effective and overall quality control of HXZQOL is currently limited due to its complex composition, especially the large amount of volatile and non-volatile active components involved. In this study, aimed to fully develop a comprehensive strategy based on non-targeted multicomponent identification, targeted authentication and quantitative analysis for quality evaluation of HXZQOL from different batches. Firstly, the non-targeted high-definition MSE (HDMSE) approach is established based on UHPLC/IM-QTOF-MS, utilized for multicomponent comprehensive characterization of HXZQOL. Combined with in house library-driven automated peak annotation and comparison of 47 reference compounds, 195 components were initially identified. In addition, HS-SPME-GC-MS was employed to analyze the volatile organic compounds (VOCs) in HXZQOL, and a total of 61 components were identified by comparison to the NIST database, reference compounds as well as retention indices. Secondly, based on the selective ion monitoring (SIM) of 24 "identity markers" (involving each herbal medicine), characteristic chromatograms (CCs) were established on LC-MS and GC-MS respectively, to authenticate 15 batches of HXZQOL samples. The targeted-SIM CCs showed that all marker compounds in 15 batches of samples could be accurately monitored, which could indicate preparations authenticity. Finally, a parallel reaction monitoring (PRM) method was established and validated to quantify the nine compounds in 15 batches of HXZQOL. Conclusively, this study first reports chemical-material basis, SIM CCs and quality evaluation of HXZQOL, which is of great implication to quality control and ensuring the authenticity of the preparation.

Authentication of pure and adulterated edible oils using non-destructive ultrasound.

At present, the quality of edible oil is evaluated using traditional analysis techniques that are generally destructive. Therefore, efforts are being made to find alternative methods with non-destructive techniques such as Ultrasound. This work aims to confirm the feasibility of non-destructive ultrasonic inspection to characterise and detect fraudulent practices in olive oil due to adulteration with two other edible vegetable oils (sunflower and corn). For this purpose, pulsed ultrasonic signals with a frequency of 2.25 MHz have been used. The samples of pure olive oil were adulterated with the other two in variable percentages between 20% and 80%. Moreover, the viscosity and density values were measured. Both these physicochemical and acoustic parameters were obtained at 24 °C and 30 °C and linearly correlated with each other. The results indicate the sensitivity of the method at all levels of adulteration studied. The responses obtained through the parameters related to the components of velocity, attenuation, and frequency of the ultrasonic waves are complementary to each other. This allows concluding that the classification of pure and adulterated oil samples is possible through non-destructive ultrasonic inspection.

Genetic Authentication of the Medicinal Plant Portulaca oleracea Using a Quick, Precise, and Sensitive Isothermal DNA Amplification Assay.

Portulaca oleracea (PO) is a commonly known medicinal crop that is an important ingredient for traditional Chinese medicine (TCM) due to its use as a vegetable in the diet. PO has been recorded to be frequently adulterated by other related species in the market of herbal plants, distorting the PO plant identity. Thus, identification of the botanical origin of PO is a crucial step before pharmaceutical or functional food application. In this research, a quick assay named "loop-mediated isothermal amplification (LAMP)" was built for the specific and sensitive authentication of PO DNA. On the basis of the divergences in the internal transcribed spacer 2 (ITS2) sequence between PO and its adulterant species, the LAMP primers were designed and verified their specificity, sensitivity, and application for the PO DNA authentication. The detection limit of the LAMP assay for PO DNA identification specifically was 100 fg under isothermal conditions at 63 °C for 30 min. In addition, different heat-processed PO samples can be applied for use in PO authentication in the LAMP assay. These samples of PO were more susceptible to the effect of steaming in authentication by PCR than boiling and drying treatment. Furthermore, commercial PO samples pursued from herbal markets were used to display their applicability of the developed LAMP analysis for PO postharvest authentication, and the investigation found that approximately 68.4% of PO specimens in the marketplace of herbal remedies were adulterated. In summary, the specific, sensitive, and rapid LAMP assay for PO authentication was first successfully developed herein, and its practical application for the inspection of adulteration in PO samples from the herbal market was shown. This LAMP assay created in this study will be useful to authenticate the botanical origin of PO and its commercial products.

Nomenclature: Herbal Taxonomy in the Global Commerce of Botanicals.

In the world trade of medicinal plants, the naming of plants is fundamental to understanding which species are acceptable for therapeutic use. There are a variety of nomenclatural systems that are used, inclusive of common names, Latinized binomials, Galenic or pharmaceutical names, and pharmacopeial definitions. Latinized binomials are the primary system used for naming wild plants, but these alone do not adequately define medicinal plant parts. Each system has its specific applications, advantages, and disadvantages. The topic of medicinal plant nomenclature is discussed broadly by underscoring when and how varying nomenclatural systems should be used. It is emphasized that pharmacopeial definitions represent the only naming system that integrates plant identity, relevant plant parts, and the specific quality metrics to which a material must comply, thus affording the most appropriate identification method available for medicinal plant materials.

Authentication and identification of Lamiaceae family with cyclic voltammetry fingerprint-PCA-LDA and determination of the used phenolic contents for classification using chromatographic analyses.

Todays, it is essential to evaluate and check the quality of herbal medicines in to protect the public health. As medicinal plants, the extracts of labiate herbs are used directly or indirectly to treat a variety of diseases. Increase in their consumption has led to the fraud in herbal medicines. Hence, modern accurate diagnostic methods must be introduced to differentiate and authenticate these samples. Electrochemical fingerprints have not been evaluated for their capacity to distinguish and classify various genera within a family. Since it is essential to classify, identify, and distinguish between these closely related plants in order to guarantee the quality of the raw materials, the authenticity and quality of 48 dried and fresh Lamiaceae samples, which include Mint, Thyme, Oregano, Satureja, Basil, and Lavender with various geographic origins, were examined. The present study focused on (a) classification and authentication Labiate herbs extracts and (b) identification of active compounds in samples by Gas chromatography and HPLC methods. This was accomplished using principal component analysis (PCA) and PCA-linear discriminate analysis (PCA-LDA). The results of the clustering revealed that PCA-LDA categorized mint species more accurately than PCA. In addition to certain flavonoids including ferulic acid, apigenin, luteolin, and quercetin, HPLC and GC analysis of the ethanolic extract revealed the presence of phenolic acids such as rosmarenic acid, methyl rosmarenate, caffeic acid, cinnamic acid, and chlorogenic acid. Comparing results of PCA-LDA with chromatographic analysis show that the authentication and detection of fraud samples were correctly performed using chemometyrics technique based on CV fingerprints. Even, there was no need to completely identify components of the mint samples.

A comparative evaluation of low-field and high-field NMR untargeted analysis: Authentication of virgin coconut oil adulterated with refined coconut oil as a case study.

Despite the advances in low-field nuclear magnetic resonance (NMR), there are limited spectroscopic applications for untargeted analysis and metabolomics. To evaluate its potential, we combined high-field and low-field NMR with chemometrics for the differentiation between virgin and refined coconut oil and for the detection of adulteration in blended samples. Although low-field NMR has less spectral resolution and sensitivity compared to high-field NMR, it was still able to achieve a differentiation between virgin and refined coconut oils, as well as between virgin coconut oil and blends, using principal component analysis (PCA), partial least squares discriminant analysis (PLS-DA), and random forest techniques. These techniques were not able to distinguish between blends with different levels of adulteration; however, partial least squares regression (PLSR) enabled the quantification of adulteration levels for both NMR approaches. Given the significant benefits of low-field NMR, including economic and user-friendly analysis and fitting in an industrial environment, this study establishes the proof of concept for its utilization in the challenging scenario of coconut oil authentication. Also, this method has the potential to be used for other similar applications that involve untargeted analysis.

Portable near-infrared (NIR) spectrometer and chemometrics for rapid identification of butter cheese adulteration.

Artisanal cheeses are highly valued around the world for their distinct sensory characteristics, thus being prone to adulteration by substituting authentic material for cheaper products, such as vegetable oil. In this work, we developed a method based on a portable NIR spectrometer as a non-destructive and low-cost alternative to identify adulteration in butter cheese. Dataset consisted of authentic and intentionally adulterated cheeses in the laboratory and commercial cheeses, which were identified as authentic and adulterated with vegetable oil after analysis of the fatty acid profile. PLS-DA classification models identified adulterated samples with an accuracy of 94.44%. PLS prediction models showed excellent performance (RPD > 3.0) to predict the adulterant level. These results demonstrate that NIR spectra can be used to identify the replacement of authentic fat by soybean oil in butter cheese and that the developed models can be used to identify adulteration in external samples with good performance.

Development and authentication of Panax ginseng cv. Sunhong with high yield and multiple tolerance to heat damage, rusty roots and lodging.

Ginseng (Panax ginseng) has been used as a valuable medicinal plant in Asia, and the demand for ginseng production for health functional food is increasing worldwide after the COVID-19 crisis. Although a number of cultivars have been developed to increase ginseng production, none of them were widely cultivated in Korea because they could not resist various environmental stresses while being grown in one place for at least 4 years. To address this, Sunhong was developed as a ginseng cultivar with high yield and multiple stress tolerance by pure line selection. Sunhong showed high yield and heat tolerance comparable to Yunpoong, a representative high-yielding cultivar, and exhibited 1.4 times lower prevalence of rusty roots than Yunpoong, suggesting that Sunhong can keep its high yield and quality during long-term cultivation. In addition, distinct color and lodging resistance were expected to increase the convenience of cultivation. To supply pure seeds to farmers, we also established a reliable high-throughput authentication system for Sunhong and seven ginseng cultivars through genotyping-by-sequencing (GBS) analysis. The GBS approach enabled to identify a sufficient number of informative SNPs in ginseng, a heterozygous and polyploid species. These results contribute to the improvement of yield, quality, and homogeneity, and therefore promote the ginseng industry. Supplementary Information: The online version contains supplementary material available at 10.1007/s13580-023-00526-x.