Variation in Botanical Reference Materials: Similarity of Actaea Racemosa Analyzed by Flow Injection Mass Spectrometry.

BACKGROUND: Botanical reference materials (BRMs) generally account for the species, cultivar, and year and location of harvest that result in variability in the chemical composition that may lead to statistically significant differences using chemometric methods. OBJECTIVE: To compare the chemical composition of five species of Actaea root BRMs, four herbal sources of A. racemosa root BRMs, and A. racemosa BRMS, and commercial roots and supplements using chemometric methods and selected pre-processing approaches. METHOD: Samples were analyzed by flow injection mass spectrometry (FIMS), principal component analysis (PCA), and factorial multivariate analysis of variance (mANOVA). RESULTS: Statistically significant (P = 0.05) compositional differences were found between three genera (Actaea, Panax, and Ginkgo), five species of Actaea (A. racemosa, A. cimicifuga, A. dahurica, A. pachypoda, and A. rubra) root BRMs, four herbal sources of A. racemosa root BRMs, and A. racemosa BRMS and commercial roots and supplements. The variability of 6% of the BRM variables was found to be quantitatively conserved and reduced the compositional differences between the four sources of root BRMs. Compositional overlap of A. racemosa and other Actaea BRMs was influenced by variation in technical repeats, pre-processing methods, selection of variables, and selection of confidence limits. Sensitivity ranged from 94 to 97% and specificity ranged from 21 to 89% for the pre-processing protocols tested. CONCLUSIONS: Environmental, genetic, and chemometric factors can influence discrimination between species and authentic botanical reference materials. HIGHLIGHTS: Frequency distribution plots derived from soft independent modeling of class analogy provide excellent means for understanding the impact of experimental factors.

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.

Chemical characterization and quantitative determination of flavonoids and phenolic acids in yerba santa (Eriodictyon spp.) using UHPLC/DAD/Q-ToF.

Eriodictyon species, commonly known as yerba santa, are plants native to the Southwestern United States and northern Mexico. The plants are known for their medicinal properties and have been used by indigenous people for centuries to treat various ailments, in particular, respiratory conditions. Despite a long history of traditional use, many of the species have never been fully chemically characterized, and the constituent range of the species has not been comprehensively reported. In an effort to establish a quality control and chemical characterization method, an extensive set of Eriodictyon species samples including E. californicum (n = 85), E. angustifolium (n = 8), E. trichocalyx (n = 5), E. crassifolium (n = 9), E. tomentosum (n = 2), E. traskiae (n = 1), and E. capitatum (n = 1) were investigated. Fourteen compounds comprised of flavonoids and phenolic acids were quantified utilizing an UHPLC/DAD method. The results from the method validation demonstrated excellent linearity (R2 > 0.99) and sensitivity as evidenced by LOD (0.01-0.1 µg/mL) and LOQ (0.05-0.2 µg/mL). Likewise, the method was found to be precise (RSD < 2.78%) with recoveries between 88.9% and 103.2%. Furthermore, by using UHPLC/ESI/Q-ToF data and protonated, deprotonated, and adduct and fragment ions in positive and negative ion modes, we were able to identify 53 compounds in yerba santa plant samples. To the best of our knowledge, this work encapsulates the most comprehensive data set currently available for the chemical characterization and quantification of the primary constituents in Eriodictyon species. Additionally, results of this study also demonstrated the applicability of the developed method for quality assessment of raw material and commercial herbal products containing different Eriodictyon species.

Botanical Ingredient Forensics: Detection of Attempts to Deceive Commonly Used Analytical Methods for Authenticating Herbal Dietary and Food Ingredients and Supplements.

Botanical ingredients are used widely in phytomedicines, dietary/food supplements, functional foods, and cosmetics. Products containing botanical ingredients are popular among many consumers and, in the case of herbal medicines, health professionals worldwide. Government regulatory agencies have set standards (collectively referred to as current Good Manufacturing Practices, cGMPs) with which suppliers and manufacturers must comply. One of the basic requirements is the need to establish the proper identity of crude botanicals in whole, cut, or powdered form, as well as botanical extracts and essential oils. Despite the legal obligation to ensure their authenticity, published reports show that a portion of these botanical ingredients and products are adulterated. Most often, such adulteration is carried out for financial gain, where ingredients are intentionally substituted, diluted, or "fortified" with undisclosed lower-cost ingredients. While some of the adulteration is easily detected with simple laboratory assays, the adulterators frequently use sophisticated schemes to mimic the visual aspects and chemical composition of the labeled botanical ingredient in order to deceive the analytical methods that are used for authentication. This review surveys the commonly used approaches for botanical ingredient adulteration and discusses appropriate test methods for the detection of fraud based on publications by the ABC-AHP-NCNPR Botanical Adulterants Prevention Program, a large-scale international program to inform various stakeholders about ingredient and product adulteration. Botanical ingredients at risk of adulteration include, but are not limited to, the essential oils of lavender (Lavandula angustifolia, Lamiaceae), rose (Rosa damascena, Rosaceae), sandalwood (Santalum album, Santalaceae), and tea tree (Melaleuca alternifolia, Myrtaceae), plus the extracts of bilberry (Vaccinium myrtillus, Ericaceae) fruit, cranberry (Vaccinium macrocarpon, Ericaceae) fruit, elder (Sambucus nigra, Viburnaceae) berry, eleuthero (Eleutherococcus senticosus, Araliaceae) root, ginkgo (Ginkgo biloba, Ginkgoaceae) leaf, grape (Vitis vinifera, Vitaceae) seed, saw palmetto (Serenoa repens, Arecaceae) fruit, St. John's wort (Hypericum perforatum, Hypericaceae) herb, and turmeric (Curcuma longa, Zingiberaceae) root/rhizome, among numerous others.

Pharmacopeial Standards for the Quality Control of Botanical Dietary Supplements in the United States.

Botanicals are among the fastest growing segments of the dietary supplement industry in the U.S. The Dietary Supplement Health and Education Act (DSHEA; Public Law 103-417 [Oct. 25, 1994]) provided a regulatory classification for the trade of numerous botanicals and botanically-derived products as dietary supplements. The global supply chain, the adoption of many botanicals that are also recognized as traditional medicines around the world as dietary supplement ingredients, and the differing recognition of the national and international pharmacopeias as sources for voluntary or mandatory quality standards present challenges in ensuring the quality of the ingredients and products. The complexity of quality assurance by compliance with pharmacopeial standards is illustrated in this article with a brief history of pharmacopoeias including their official recognition in national laws, their approaches to the science behind the standards, the use of reference standards for quality assessment and regulatory compliance, the use of pharmacopeial standards by the industry and regulators within the DSHEA framework in the United States, and a discussion of the global supply chain. Pharmacopeial standards can help regulators and the industry adapt to the new technologies that present both opportunities and challenges.

Development of a Metabolite Ratio Rule-Based Method for Automated Metabolite Profiling and Species Differentiation of Four Major Cinnamon Species.

A metabolomic ratio rule-based classification method was developed and programmed for automated metabolite profiling and differentiation of four major cinnamon species using ultra-high-performance liquid chromatography-high-resolution mass spectrometry (UHPLC-HRMS). The computational program identifies key cinnamon metabolites, including proanthocyanidins, cinnamaldehyde, and coumarin, from test samples through LC-MS data processing and assigns cinnamon species by critical metabolite ratios using a stepwise classification strategy. Further, 100% classification accuracy was achieved on the training sample set through critical ratio optimization, and over 95% accuracy was achieved on the validation sample set. The proposed cinnamon classification method exhibited superior accuracy compared to the metabolomic-based PLS-DA modeling method and offered great value for the authentication of cinnamon samples and evaluation of their potential health benefits.

Potentially toxic pyrrolizidine alkaloids in Eupatorium perfoliatum and three related species. Implications for herbal use as boneset.

INTRODUCTION: Pro-toxic dehydropyrrolizidine alkaloids are associated with liver disease in humans. The potential for long-term, low-level or intermittent exposures to cause or contribute to chronically-developing diseases is of international concern. Eupatorium perfoliatum is a medicinal herb referred to as boneset. While the presence of dehydropyrrolizidine alkaloids in some Eupatorium species is well-established, reports on Eupatorium perfoliatum are scant and contradictory. OBJECTIVE: To investigate the presence of dehydropyrrolizidine alkaloids in a survey of boneset samples and related alcoholic tinctures, and hot water infusions and decoctions. METHODS: Methanol, hot water or aqueous ethanol extracts of Eupatorium perfoliatum and three closely-related species were subjected to HPLC-ESI(+)MS and MS/MS analysis using three complementary column methods. Dehydropyrrolizidine alkaloids were identified from their MS data and comparison with standards. RESULTS: Forty-nine samples of Eupatorium perfoliatum were shown to contain dehydropyrrolizidine alkaloids (0.0002-0.07% w/w), the majority dominated by lycopsamine and intermedine, their N-oxides and acetylated derivatives. Alcoholic tinctures and hot water infusions and decoctions had high concentrations of the alkaloids. Different chemotypes, hybridisation or contamination of some Eupatorium perfoliatum samples with related species were suggested by the co-presence of retronecine- and heliotridine-based alkaloids. CONCLUSIONS: Sampling issues, low and high alkaloid chemotypes of Eupatorium perfoliatum or interspecies hybridization could cause the wide variation in dehydropyrrolizidine alkaloid concentrations or the different profiles observed. Concerns associated with dehydropyrrolizidine alkaloids provide a compelling reason for preclusive caution until further research can better define the toxicity and carcinogenicity of the dehydropyrrolizidine alkaloid content of Eupatorium perfoliatum. [Correction added on 12 July 2018, after first online publication: The 'Conclusions' section in the abstract has been added.].

Identification of Ginkgo biloba supplements adulteration using high performance thin layer chromatography and ultra high performance liquid chromatography-diode array detector-quadrupole time of flight-mass spectrometry.

Ginkgo biloba is one of the most widely sold herbal supplements and medicines in the world. Its popularity stems from having a positive effect on memory and the circulatory system in clinical studies. As ginkgo popularity increased, non-proprietary extracts were introduced claiming to have a similar phytochemical profile as the clinically tested extracts. The standardized commercial extracts of G. biloba leaf used in ginkgo supplements contain not less than 6% sesquiterpene lactones and 24% flavonol glycosides. While sesquiterpene lactones are unique constituents of ginkgo leaf, the flavonol glycosides are found in many other botanical extracts. Being a high value botanical, low quality ginkgo extracts may be subjected to adulteration with flavonoids to meet the requirement of 24% flavonol glycosides. Chemical analysis by ultra high performance liquid chromatography-mass spectrometry revealed that adulteration of ginkgo leaf extracts in many of these products is common, the naturally flavonol glycoside-rich extract being spiked with pure flavonoids or extracts made from another flavonoid-rich material, such as the fruit/flower of Japanese sophora (Styphnolobium japonicum), which also contains the isoflavone genistein. Recently, genistein has been proposed as an analytical marker for the detection of adulteration of ginkgo extracts with S. japonicum. This study confirms that botanically authenticated G. biloba leaf and extracts made therefrom do not contain genistein, and the presence of which even in trace amounts is suggestive of adulteration. In addition to the mass spectrometric approach, a high performance thin layer chromatography method was developed as a fast and economic method for chemical fingerprint analysis of ginkgo samples.

Hypericum perforatum: pharmacokinetic, mechanism of action, tolerability, and clinical drug-drug interactions.

Hypericum perforatum (HP) belongs to the Hypericaceae family and is one of the oldest used and most extensively investigated medicinal herbs. The medicinal form comprises the leaves and flowering tops of which the primary ingredients of interest are naphthodianthrones, xanthones, flavonoids, phloroglucinols (e.g. hyperforin), and hypericin. Although several constituents elicit pharmacological effects that are consistent with HP's antidepressant activity, no single mechanism of action underlying these effects has thus far been found. Various clinical trials have shown that HP has a comparable antidepressant efficacy as some currently used antidepressant drugs in the treatment of mild/moderate depression. Interestingly, low-hyperforin-content preparations are effective in the treatment of depression. Moreover, HP is also used to treat certain forms of anxiety. However, HP can induce various cytochrome P450s isozymes and/or P-glycoprotein, of which many drugs are substrates and which are the main origin of HP-drug interactions. Here, we analyse the existing evidence describing the clinical consequence of HP-drug interactions. Although some of the reported interactions are based on findings from in vitro studies, the clinical importance of which remain to be demonstrated, others are based on case reports where causality can, in some cases, be determined to reveal clinically significant interactions that suggest caution, consideration, and disclosure of potential interactions prior to informed use of HP.

Use of high-performance thin layer chromatography by the American Herbal Pharmacopoeia.

TLC characterizations are among the key identity tests in most pharmacopoeial monographs. Pharmacopoeial standards are typically used by industry as a basis for meeting QC requirements and current good manufacturing practices (cGMPs). TLC is a relatively low-cost, highly versatile tool for developing specifications for raw materials, as well as for the various preparations for which pharmacopoeial standards are created. In addition to its use in the development of identity tests, TLC is a valuable tool for screening plant samples that pharmacopoeias must review in the development of monographs and botanical reference materials (BRMs). Specifically, HPTLC is the ideal TLC technique for these purposes because of its increased accuracy, reproducibility, and ability to document the results, compared with standard TLC. Because of this, HPTLC technologies are also the most appropriate TLC technique for conformity with GMPs. This article highlights the manner in which HPTLC is used by the American Herbal Pharmacopoeia (AHP) in the development of AHP monograph identity standards, the identification of adulterating species, and the development of AHP-verified BRMs.

Comparison of the phenolic component profiles of skullcap (Scutellaria lateriflora) and germander (Teucrium canadense and T. chamaedrys), a potentially hepatotoxic adulterant.

INTRODUCTION: Scutellaria lateriflora, commonly known as skullcap, is used as an ingredient in numerous herbal products. Unfortunately, it has occasionally been adulterated with Teucrium canadense or T. chamaedrys, commonly known as germander, which contains potentially hepatotoxic diterpenes. Chromatographic profiles of the phenolic components provide a means of distinguishing between these plants and enhancing public safety. OBJECTIVE: To develop a chromatographic method for the identification of Scutellaria lateriflora and two Teucrium species and to quantify the latter as adulterants. METHODOLOGY: Samples were extracted with aqueous methanol and the extracts were analysed using a standardised LC-DAD-ESI/MS profiling method to obtain their phenolic profiles. RESULTS: Skullcap contained primarily flavonoids, while the major phenolic components of the two Teucrium species were the phenylethanoids, verbascoside and teucrioside. Using the phenylethanoids as markers, it was possible to clearly distinguish between the two genus and to determine 5% Teucrium mixed with Scutellaria using either ultraviolet absorption spectrometry or mass spectrometry in the total ion count mode. Using MS in the selective ion monitoring (SIM) mode, 1% Teucrium could be measured. CONCLUSIONS: This study showed that chromatographic profiling was able to identify Scutellaria and Teucrium, separately and when mixed together.

Chromatographic fingerprint analysis--a rational approach for quality assessment of traditional Chinese herbal medicine.

Traditional Chinese Herbal Medicine (TCHM) contain multiple botanicals, each of which contains many compounds that may be relevant to the medicine's putative activity. Therefore, analytical techniques that look at a suite of compounds, including their respective ratios, provide a more rational approach to the authentication and quality assessment of TCHM. In this paper we present several examples of applying chromatographic fingerprint analysis for determining the identity, stability, and consistency of TCHM as well as the identification of adulterants as follows: (1) species authentication of various species of ginseng (Panax ginseng, Panax quinquefolium, Panax noto-ginseng) and stability of ginseng preparations using high performance thin-layer chromatography (HPTLC) fingerprint analysis; (2) batch-to-batch consistency of extracts of Total Glycosides of Peony (TGP), to be used as a raw material and in finished products (TGP powdered extract products), using high performance liquid chromatography (HPLC) fingerprint analysis with a pattern recognition software interface (CASE); (3) documenting the representative HPLC fingerprints of Immature Fruits of Terminalia chebula (IFTC) through the assessment of raw material, in-process assay of the extracts, and the analysis of the finished product (tablets); (4) HPLC fingerprint study demonstrating the consistent quality of total flavonoids of commercial extracts of ginkgo (Ginkgo biloba) leaves (EGb) along with detection of adulterations. The experimental conditions as well as general comments on the application of chromatographic fingerprint analysis are discussed.