6-Oxofurostane and (iso)Spirostane Types of Saponins in Smilax sieboldii: UHPLC-QToF-MS/MS and GNPS-Molecular Networking Approach for the Rapid Dereplication and Biodistribution of Specialized Metabolites.

Identifying novel phytochemical secondary metabolites following classical pharmacognostic investigations is tedious and often involves repetitive chromatographic efforts. During the past decade, Ultra-High Performance Liquid Chromatography-Quadrupole Time of Flight-Tandem Mass Spectrometry (UHPLC-QToF-MS/MS), in combination with molecular networking, has been successfully demonstrated for the rapid dereplication of novel natural products in complex mixtures. As a logical application of such innovative tools in botanical research, more than 40 unique 3-oxy-, 3, 6-dioxy-, and 3, 6, 27-trioxy-steroidal saponins were identified in aerial parts and rhizomes of botanically verified Smilax sieboldii. Tandem mass diagnostic fragmentation patterns of aglycones, diosgenin, sarsasapogenin/tigogenin, or laxogenin were critical to establishing the unique nodes belonging to six groups of nineteen unknown steroidal saponins identified in S. sieboldii. Mass fragmentation analysis resulted in the identification of 6-hydroxy sapogenins, believed to be key precursors in the biogenesis of characteristic smilaxins and sieboldins, along with other saponins identified within S. sieboldii. These analytes' relative biodistribution and characteristic molecular networking profiles were established by analyzing the leaf, stem, and root/rhizome of S. sieboldii. Deducing such profiles is anticipated to aid the overall product integrity of botanical dietary supplements while avoiding tedious pharmacognostic investigations and helping identify exogenous components within the finished products.

Daphne jejudoensis Attenuates LPS-Induced Inflammation by Inhibiting TNF-α, IL-1ß, IL-6, iNOS, and COX-2 Expression in Periodontal Ligament Cells.

Periodontitis is a common disease involving inflammation and tissue destruction in the periodontal region. Although uncontrolled long-term inflammation in the gingiva may lead to loss of the periodontal ligament, treatments or preventive solutions for periodontitis are scarce. The aim of this study is to find anti-inflammatory material from a natural source that can be used to treat or protect against periodontitis. Daphne species (Thymelaeaceae) are important and popular components of traditional Chinese medicine and are used as anti-inflammatory agents. Daphne jejudoensis is an endemic plant that grows on Jeju Island and was identified as a new species in 2013. In this study, for the first time, we investigated the anti-inflammatory effect of D. jejudoensis leaf extract (DJLE) on human periodontal ligament cells. The gene expression levels of pro-inflammatory cytokines (interleukin-1ß and 6 and tumor necrosis factor-α) and inflammation-inducible enzymes (inducible nitric oxide synthase and cyclooxygenase-2) were reduced after DJLE treatment with/without lipopolysaccharide stimulation. The findings of this study indicate that D. jejudoensis possesses anti-inflammatory activities, suggesting that DJLE may be a potential preventive and therapeutic agent for periodontitis.

A Comprehensive Workflow for the Analysis of Bio-Macromolecular Supplements: Case Study of 20 Whey Protein Products.

The presence of bio-macromolecules as major ingredients is a primary factor in marketing many biologically derived macromolecular supplements. Workflows for analyzing these supplements for quality assurance, adulteration, and other supply-chain difficulties must include a qualitative assessment of small-molecule and macromolecular components; however, no such integrated protocol has been reported for these bio-macromolecular supplements. Twenty whey protein supplements were analyzed using an integrated workflow to identify protein content, protein adulteration, inorganic elemental content, and macromolecular and small-molecule profiles. Orthogonal analytical methods were employed, including NMR profiling, LC-DAD-QToF analysis of small-molecule components, ICP-MS analysis of inorganic elements, determination of total protein content by a Bradford assay, SDS-PAGE protein profiling, and bottom-up shotgun proteomic analysis using LC-MS-MS. All 20 supplements showed a reduced protein content compared to the claimed content but no evidence of adulteration with protein from an unclaimed source. Many supplements included unlabeled small-molecule additives (but nontoxic) and significant deviations in metal content, highlighting the importance of both macromolecular and small-molecule analysis in the comprehensive profiling of macromolecular supplements. An orthogonal, integrated workflow allowed the detection of crucial product characteristics that would have remained unidentified using traditional workflows involving either analysis of small-molecule nutritional supplements or protein analysis.

Chemical Profiling and Characterization of Anthraquinones from Two Bulbine Species and Dietary Supplements Using Liquid Chromatography-High Resolution Mass Spectrometry.

BACKGROUND: Bulbine natalensis Baker and Bulbine frutescens (L.) Willd., belonging to the family Asphodelaceae, are widely distributed in South Africa and traditionally used as an aphrodisiac and skin remedies. OBJECTIVE: The aim of this study is to develop an analytical method for chemical profiling and identification of components in Bulbine species, which would be useful for herbal identification and understanding of the biological activity of B. natalensis in terms of safety and benefits to human health. METHOD: The anthraquinone-type compounds were structurally characterized from the extracts of dried stem and roots of Bulbine species and dietary supplements using liquid chromatography-quadrupole time-of-flight mass spectrometry (LC-QToF) with negative and positive ion electrospray. The calculated accurate masses of the protonated and deprotonated molecules and fragment ions were used for identification of the components from two Bulbine species. RESULTS: A total of 55 anthraquinone-type compounds, including 11 standard compounds, were identified in the crude extracts of two Bulbine species. Two Bulbine species and dietary supplements were clustered into different groups and possible chemical markers were identified. CONCLUSIONS: The developed analytical method provided a fast and economic method for quality assessment of Bulbine species in dietary supplements based on anthraquinone-type compounds. HIGHLIGHTS: This study reports holistic chemical profiling of Bulbine species using LC-QToF. The developed analytical method enabled non-targeted analysis of components in B. natalensis and B. frutescens, and is recommended for commercial and regulatory purposes.

Quantitative determination and characterization of polyphenols from Cissus quadrangularis L. and dietary supplements using UHPLC-PDA-MS, LC-QToF and HPTLC.

Stem and leaf of Cissus quadrangularis L. (Vitaceae), indigenous to Asia and Africa, were used for medicinal and dietary purposes with limited information about the plant's phytochemistry. Stem and leaf samples were assessed for the simultaneous determination of polyphenolic compounds (catechin, epicatechin, quercetin-3-O-ß-glucopyranoside, kaempferol-3-O-ß-glucoside, quercetin-3-O-ß-rhamnoside, leachianol F, amurensin A, pallidol, resveratrol, and quadrangularin A), using UHPLC-PDA-MS. The validation data showed that the method is precise, specific, accurate, and linear over the range of 0.5-100 µg/mL. Reversed-phase ultra-high performance liquid chromatography (UHPLC) fingerprints of the crude methanolic stem and leaf extracts of C. quadrangularis were obtained at different wavelengths based on their λmax. Polyphenolics were characterized using both UHPLC-PDA-MS and LC-QToF analysis. From liquid chromatography quadrupole time of flight-electrospray ionization mass spectrometry (LC-QToF) spectra, over 40 components were structurally correlated, and confirmation was based on the fragmentation characteristics and also from the information available in the literature. In addition to the LC-QToF method, a simple, fast HPTLC method was developed as a visual aid for the rapid qualitative analytical tool to help establish the quality assessment of botanical raw materials and dietary supplements claiming to contain Cissus.

Dibenzocyclooctadiene Lignans in Plant Parts and Fermented Beverages of Schisandra chinensis.

The fruit of Schisandra chinensis, Omija, is a well-known traditional medicine used as an anti-tussive and anti-diarrhea agent, with various biological activities derived from the dibenzocyclooctadiene-type lignans. A high-pressure liquid chromatography-diode array detector (HPLC-DAD) method was used to determine seven lignans (schisandrol A and B, tigloylgomisin H, angeloylgomisin H, schisandrin A, B, and C) in the different plant parts and beverages of the fruit of S. chinensis grown in Korea. The contents of these lignans in the plant parts descended in the following order: seeds, flowers, leaves, pulp, and stems. The total lignan content in Omija beverages fermented with white sugar for 12 months increased by 2.6-fold. Omija was fermented for 12 months with white sugar, brown sugar, and oligosaccharide/white sugar (1:1, w/w). The total lignan content in Omija fermented with oligosaccharide/white sugar was approximately 1.2- and 1.7-fold higher than those fermented with white sugar and brown sugar, respectively. A drink prepared by immersion of the fruit in alcohol had a higher total lignan content than these fermented beverages. This is the first report documenting the quantitative changes in dibenzocyclooctadiene-type lignans over a fermentation period and the effects of the fermentable sugars on this eco-friendly fermentation process.

Modulation of CYP3A4 and CYP2C9 activity by Bulbine natalensis and its constituents: An assessment of HDI risk of B. natalensis containing supplements.

BACKGROUND: Bulbine natalensis is an African-folk medicinal plant used as a dietary supplement for enhancing sexual function and muscle strength in males by presumably boosting testosterone levels, but no scientific information is available about the possible herb-drug interaction (HDI) risk when bulbine-containing supplements are concomitantly taken with prescription drugs. PURPOSE: This study was aimed to investigate the HDI potential of B. natalensis in terms of the pregnane X receptor (PXR)-mediated induction of major drug-metabolizing cytochrome P450 enzyme isoforms (i.e., CYP3A4 and CYP2C9) as well as inhibition of their catalytic activity. RESULTS: We found that a methanolic extract of B. natalensis activated PXR (EC50 6.2 ± 0.6 µg/ml) in HepG2 cells resulting in increased mRNA expression of CYP3A4 (2.40 ± 0.01 fold) and CYP2C9 (3.37 ± 0.3 fold) at 30 µg/ml which was reflected in increased activites of the two enzymes. Among the constituents of B. natalensis, knipholone was the most potent PXR activator (EC50 0.3 ± 0.1 µM) followed by bulbine-knipholone (EC50 2.0 ± 0.5 µM), and 6'-methylknipholone (EC50 4.0 ± 0.5 µM). Knipholone was also the most effective in increasing the expression of CYP3A4 (8.47 ± 2.5 fold) and CYP2C9 (2.64 ± 0.3 fold) at 10 µM. Docking studies further confirmed the unique structural features associated with knipholones for their superior inductive potentials in the activation of PXR compared to other anthraquinones. In a CYP inhibition assay, the methanolic extract as well as the anthraquinones strongly inhibited the catalytic activity of CYP2C9 while, inhibition of CYP3A4 was weak. CONCLUSIONS: These results suggest that consumption of B. natalensis may pose a potential risk for HDI if taken with conventional medications that are substrates of CYP3A4 and CYP2C9 and may contribute to unanticipated adverse reactions or therapeutic failures. Further studies are warranted to validate these findings and establish their clinical relevancy.

Quantification and Characterization of Phenolic Compounds from Northern Indian Propolis Extracts and Dietary Supplements.

BACKGROUND: Propolis is a resinous substance produced by bees. Propolis extracts have been used for anti-inflammatory and antimicrobial activities. The use of propolis dietary supplements has been increasing in the United States and the rest of the world. OBJECTIVE: A simple, economic, and valid analytical method is needed for quality assessment of dietary supplements and extracts claiming to contain propolis. METHODS: A ultra-high performance liquid chromatography (UHPLC) quadropole time-of-flight-MS method was used to characterize the chemical composition of northern Indian propolis. Fourteen major phenolic compounds were quantified using a UHPLC-DAD method. An HPTLC method was used to develop chemical fingerprinting profiles for propolis extracts and dietary supplements. The seven propolis extracts and 14 dietary supplements purchased in the U.S. were analyzed using the UHPLC-DAD-QToF method. RESULTS: Fifty-seven compounds belonging to phenolic, coumarin, fatty acid, and terpene classes were identified in propolis extracts. Based on quantification results, the content of 14 phenolic compounds in propolis extracts varied from 19-32% in dietary supplements, a significant variation to the recommended daily intake (0.2-94 mg/day). CONCLUSIONS/HIGHLIGHTS: The developed analytical methods can be used for quality assessment of propolis extracts and dietary supplements.

Chemical profiling and characterization of phenolic acids, flavonoids, terpene glycosides from Vangueria agrestis using ultra-high-performance liquid chromatography/ion mobility quadrupole time-of-flight mass spectrometry and metabolomics approach.

Vangueria agrestis is a shrub indigenous to tropical Africa, belonging to family Rubiaceae and is traditionally used as a decoction for treatment of fever, pain, and malaria. This study was undertaken to investigate the chemical constituents based on precursor exact mass and fragment ion information. The chemical profiling and structural characteristics of chemical constituents from methanolic extracts of dried aerial parts and roots of V. agrestis and dietary supplements were analyzed using ultra-high-performance liquid chromatography/ion mobility quadrupole time-of-flight mass spectrometry coupled with UNIFI platform and multivariate analysis in both negative and positive ion modes. A non-targeted ultra-high-performance liquid chromatography-mass spectrometry analysis was carried out to profile the chemical constituents of crude extracts of V. agrestis, and 73 compounds, including reference compounds, were identified. The fragments of flavonoids, monoterpene, and triterpene glycosides revealed the characteristic cleavage of glycosidic linkages, and the fragmentation pattern provided the identity of the sugars. This analytical method provides a quick method for quality assessment of dietary supplements. Finally, a chemometrics approach with multivariate statistical tools was used to visualize the differences between root and aerial parts of plant samples and to find the potential chemical markers that differentiate among these parts of V. agrestis samples and dietary supplements.

The scoop on brain health dietary supplement products containing huperzine A.

Context: Public health concerns are emerging surrounding huperzine A commonly found in dietary supplements. We sought to determine the actual content of products claiming to contain huperzine A and whether the ingredients on the supplement facts labels matched the analyses.Methods: We identified and analyzed 22 dietary supplement products listing huperzine A on product labels. We found these products were listed in Natural Medicines and Dietary Supplement Databases and being queried by Military Service Members for enhanced mental focus, alertness and energy. Analyses were conducted by using Liquid Chromatography-Quadrupole Time of Flight Mass Spectrometry.Results: Sixteen (73%) products had at least one ingredient claimed on the supplement facts label not detected through analysis. Compounds not reported on the label were detected in 16 (73%) products analyzed. Nine products (41%) listed ingredients not meeting the regulations for being a dietary supplement ingredient according to the FDA. Ingredients of most concern detected include stimulants: demelverine, 1,5-dimethylhexylamine, 1,3-dimethylhexylamine, N-phenethyl dimethylamine, halostachine, higenamine, noopept, ß-PEA, vinpocetine, sulbutiamine; and hordenine, currently on the FDA advisory list. Quantitative analysis showed the presence of huperzine A in the range from detected under the limits of quantification (DUL) to 267.1 µg/serving. Only two supplements showed huperzine A content within 10% of the declared amount.Conclusions: In a study of dietary supplements claiming to contain huperzine A, we found products that had at least one ingredient claimed on the supplement facts label not detected through analysis. Moreover, some ingredients not on the label could be dangerous and likely do not meet the definition of a dietary supplement ingredient according to the FDA. Quantitative analysis of huperzine A showed the amount detected was not in line with what appeared on the product label. Consumers should be aware of deceptive label claims and warned not to purchase products containing potentially dangerous ingredients.

Development and Validation of a UHPLC-PDA-MS Method for the Quantitative Analysis of Anthraquinones in Bulbine natalensis Extracts and Dietary Supplements.

A UHPLC-photodiode array-MS method was developed and validated for the quantification of one chromone and six anthraquinone type of compounds from Bulbine natalensis plant samples and dietary supplements. Metabolites 1:  -  7: were identified based on their retention times and electrospray ionization-MS spectra compared with a mix of previously isolated compounds. The quantification of 1:  -  7: was based on photodiode array detection. The optimized separation was achieved using a CORTECS C18 column with a gradient of water/acetonitrile as the mobile phase. Seven compounds were separated within 15 minutes with detection limits of 50 pg on the column. The analytical method was validated for linearity, repeatability, accuracy, limits of detection, and limits of quantification. The relative standard deviations for intra- and inter-day experiments were less than 5% and the recovery efficiency was 98 - 101%. Nine dietary supplements labeled as containing B. natalensis were examined. Anthraquinone-type compounds were detected in only five out of nine dietary supplements, with the total amount ranging from 11.3 to 90.4 mg per daily dose. The analytical method is simple, economic, rapid, and can be applied for quality assessment of B. natalensis and dietary supplements. Electrospray ionization-MS was used for the identification of these compounds in plant samples and dietary products.

Analysis of prenylflavonoids from aerial parts of Epimedium grandiflorum and dietary supplements using HPTLC, UHPLC-PDA and UHPLC-QToF along with chemometric tools to differentiate Epimedium species.

An UHPLC method was developed for the determination of 15 prenylflavonoids from aerial parts of Epimedium grandiflorum and related species (Berberidaceae). The separation was achieved using a reverse phased column and water/acetonitrile gradient as a mobile phase at a temperature of 40°C. The developed analytical method was validated for linearity, limits of detection (LOD) and limits of quantification (LOQ), stability and repeatability. The LOD and LOQ were found to be in the range from 0.1-0.5 µg/mL and 0.3-1 µg/mL, respectively. The wavelength used for quantification with the photodiode array detector was 269 nm. The total content of 15 prenylflavonoids was 9.1-20.6 mg/g for E. grandiflorum (except for sample #2899 and #20862), 5.6-35.4 mg/g for E. brevicornu and 10.8-30.5 mg/g for E. sagittatum. Twenty dietary supplements contained in the range from 0.1 to 81.7 mg/day. The developed method is simple, rapid and especially suitable for quality assessment of E. grandiflorum and dietary supplements containing E. grandiflorum. Liquid chromatography quadrupole time-of-flight-mass spectrometry (LC-QToF) is described for the identification and confirmation of compounds in plant samples and dietary supplements. This technique is also used for chemical profiling of Epimedium samples. This method involved the use of protonated ions in the positive ion mode and deprotonated ions in the negative ion mode with extracted ion chromatogram (EIC). Chemometric analytical tools for visualizing the plant and commercial samples quality were used for discriminating between Epimedium species and dietary supplements with regards to the relative content or presence of components. A HPTLC method was also developed for the fast chemical fingerprint analysis of Epimedium species.

Liquid chromatography-quadrupole time of flight mass spectrometric method for targeted analysis of 111 nitrogen-based compounds in weight loss and ergogenic supplements.

The use of supplements for weight loss and in sports as pre-workout (ergogenic) products is widespread. Many of these supplements were found to contain active components, which were not claimed on the products labels. A validated liquid chromatography high-resolution mass spectrometry quadrupole time-of-flight (LC-QToF-MS) method was developed for the simultaneous analysis of 111 amine-based compounds belonging to ergogenics, anorectics and other active components including phenethylamines (amphetamines, ephedrines), sibutramine or yohimbine. This method involves the detection of [M+H]+ ions and the separation was achieved using a C18 column, water/acetonitrile gradient as the mobile phase. The method was validated for linearity, repeatability, accuracy, stability, system suitability, limits of quantification (LOQ) and limits of detection (LOD). The limits of detection were in the range from 0.001-0.5 µg/mL. The validated method was applied to the analysis of twenty-seven weight loss and ergogenic dietary supplements. Two-thirds of the supplements contained compounds that were not listed on the product's label. These include several phenethylamines (PEA) such as demelverine, hordenine, N, N-dimethyl-phenethylamine, synephrine, N-methyl-ß-phenethylamine, and methylsynephrine. In addition, the PEA mimics such as dimethylamylamine, dimethylbutylamine other stimulants including fursultiamine, evodiamine, phenibut and theophylline were also observed. One or more of the ingredients listed on the labels were not detected in forty-four percent of the products analyzed. Positive identification was based on retention time, accurate mass and fragment ions in comparison with the respective reference standards. Development of such methods is anticipated to be of aid to regulatory agencies for the identification of undeclared exogenous components that are found in many dietary supplement products.

A new anti-Helicobacter pylori juglone from Reynoutria japonica.

A 70% ethanol extract from the root portion of Reynoutria japonica afforded one new and three known juglone derivatives, namely, 2-methoxy-6-acetyl-7-methyljuglone (1), 2-ethoxy-6-acetyl-7-methyljuglone (2), 2-methoxy-7-acetonyljuglone (3), and 3-acetyl-7-methoxy-2-methyljuglone (4) together with two phenolics (5 and 6), an anthraquinone (7), a stilbene (8) and a phthalide (9). Their structures were elucidated on the basis of comprehensive spectroscopic studies including IR, MS, and 1H, 13C, 2D NMR spectra. Compound 3 is a new compound in nature, and compounds 4-6 have been isolated for the first time from R. japonica. The isolates were evaluated for their antibacterial activity against three strains (43504, 51, and 26695) of Helicobacter pylori. The four isolated juglone derivatives (1-4) showed potent growth inhibitory activity. Among them, compounds 1-3 exhibited stronger inhibitory activity than those of the positive controls, juglone and metronidazole, for the three strains and that of another reference, clarithromycin, for the 43504 and 51 strains. Specifically, the new juglone compound 3 displayed the most potent antibacterial activity against all three strains, 43504, 51, and 26695, with MIC values of 0.06, 0.06 and 0.13 µM, respectively, and MIC50 values of 0.14, 0.11 and 0.15 µM, respectively.

Decaffeinated Green Tea Extract Does Not Elicit Hepatotoxic Effects and Modulates the Gut Microbiome in Lean B6C3F1 Mice.

The main purpose of this study was to investigate the hepatotoxic potential and effects on the gut microbiome of decaffeinated green tea extract (dGTE) in lean B6C3F1 mice. Gavaging dGTE over a range of 1X-10X mouse equivalent doses (MED) for up to two weeks did not elicit significant histomorphological, physiological, biochemical or molecular alterations in mouse livers. At the same time, administration of dGTE at MED comparable to those consumed by humans resulted in significant modulation of gut microflora, with increases in Akkermansia sp. being most pronounced. Results of this study demonstrate that administration of relevant-to-human-consumption MED of dGTE to non-fasting mice does not lead to hepatotoxicity. Furthermore, dGTE administered to lean mice, caused changes in gut microflora comparable to those observed in obese mice. This study provides further insight into the previously reported weight management properties of dGTE; however, future studies are needed to fully evaluate and understand this effect.

The power of hyphenated chromatography-Time of flight mass spectrometry for unequivocal identification of spirostanes in bodybuilding dietary supplements.

A previously unidentified purported botanical ingredient was found in dietary supplements marketed for anabolic benefits. In an attempt to assess the 'naturalness' of a group of steroid-like compounds called laxogenins, a UHPLC-QToF method was developed. Several dietary supplements claim to contain 5α-hydroxy laxogenin, which is a derivative of a naturally occurring spirostane-type steroid, laxogenin. Although laxogenin has been isolated from the rhizomes of Smilax sieboldii, 5α-hydroxy laxogenin has not been isolated or reported from any natural source. These derivatives of laxogenins have untested anabolic properties. Due to the low UV absorbance of the spirostanes, a mass spectrometric method in positive ion mode was developed for unambiguous identification of laxogenin and closely related compounds. To show the utility of the developed method, twelve dietary supplements labeled to contain 5α-hydroxy laxogenin or laxogenin as 5α-hydroxy laxogenin were analyzed as a proof-of-concept. Five supplements did not contain any 5α-hydroxy laxogenin, whereas in the remaining seven samples, spirostane-type contaminants were identified along with the labeled 5α-hydroxy laxogenin. The identity of some of these contaminants was established based on reference standards along with mass fragmentation patterns. One of the unlabeled contaminants was identified as the phytosteroid saponin, diosgenin, a common starting precursor of several steroidal drugs. Several synthetic derivatives of diosgenin were identified in the eight products. These findings indicate that the labeled 5α-hydroxy laxogenin along with other spirostanes found in supplements are synthetic and signify a lack of quality controls. Additionally, an unlabeled, anabolic androgenic steroid, arimistane, an aromatase inhibitor, was also identified in one product. Laxogenin, was not detected in any of the samples analyzed during this investigation.

Quantification of Phenolic Compounds from Fadogia agrestis and Dietary Supplements using UHPLC-PDA-MS.

Fadogia agrestis is used in traditional African medicine as an analgesic and for anti-inflammatory and aphrodisiac activities. An ultra-high-performance liquid chromatography method was developed for the determination of 11 chemical constituents from roots and aerial parts of F. agrestis. The separation was achieved within 7 min by using C-18 column material and a water/acetonitrile mobile phase, both containing 0.1% formic acid gradient system with a temperature of 45 °C. The method was validated for linearity, repeatability, limits of detection, and limits of quantification. The limits of detection of phenolic compounds were found to be in the range from 0.025 to 0.1 µg/mL. The wavelengths used for quantification with the photodiode array detector were 238, 254, 291, and 325 nm. Twelve of 17 dietary supplements contained phenolic compounds in the range from 0.3 to 2.7 mg/d. The phenolic compounds were not detected in five dietary supplements. Liquid chromatography-mass spectrometry coupled with electrospray ionization interface method is described for the identification and confirmation of compounds from plant samples and dietary supplements claiming to contain F. agrestis. This method involved the use of [M + H]+ and [M + Na]+ ions in the positive mode and [M - H]- ions in the negative mode with extractive ion monitoring. The developed method is simple, economic, rapid, and especially suitable for quality control and chemical fingerprint analysis of F. agrestis.

Microscopic characterization and HPTLC of the leaves, stems and roots of Fadogia agrestis - an African folk medicinal plant

ABSTRACT Fadogia agrestis Schweinf. ex Hiern (Vangueria agrestis (Schweinf. ex Hiern) Lantz), Rubiaceae, is an African traditional medicinal plant also used as a dietary supplement in the US. The present paper is the first report of the pharmacognostic study of the leaf, stem and root of F. agrestis by microscopy, HPTLC and total phenolic/flavonoid content analyses. Noteworthy microscopic features that can help in identification and quality control are septate and lignified non-glandular trichomes on leaf and stem epidermises, paracytic stomata on leaf abaxial epidermis, numerous cells containing yellow substances of presumably phenolic compounds in leaf and stem, calcium oxalate druses and prismatic crystals in leaf and styloids in stem, primary phloem fibers in stem, brachysclereids in stem and root, spherical starch grains in root, and vessels with vestured pits and simple perforated end walls. In addition to microscopy, a total phenolic/flavonoid content determination and an HPTLC method were also developed for rapid chemical fingerprint analyses of Fadogia samples and dietary supplements.

Targeted and non-targeted analysis of annonaceous alkaloids and acetogenins from Asimina and Annona species using UHPLC-QToF-MS.

In current work, targeted and non-targeted analysis of alkaloids and acetogenins from methanolic extracts of Asimina, Annona species and dietary supplements have been performed using UHPLC-QToF in positive ion mode. Thirty-five standard compounds (twelve alkaloids and twenty-three acetogenins) were used for the analysis. The fragment ions produced by collision induced dissociation (CID) revealed the characteristic cleavage and provided structural information. Aporphine alkaloids and acetogenins are the major groups found in Asimina and Annona species. An untargeted analysis based on high-resolution mass spectrometry was carried out to profile the alkaloids and acetogenins from Asimina species (As. triloba, As. parviflora). Magnoflorine, being a major alkaloid from twigs of As. triloba samples, was used as an example to discuss the fragmentation patterns. In (+)-ESI-MS, magnoflorine gave [M]+ ions at m/z 342.1705. The fragment ions at m/z 297.1127 [M-(CH3)2NH]+, 282.0886 [M-(CH3)3NH]+, 265.0865 [M-(CH3)2NH-CH3OH]+, 237.0916 [M-(CH3)2NH-CH3OH-CO]+, and 222.0681 [M-(CH3)2NH-CH3OH-CO-CH3]+ resulted from the [M]+ molecular ion. One dietary supplement claiming to contain paw paw (As. triloba) was also analyzed and showed a similar profile to twigs of As. triloba. A total of 131 compounds including standard compounds were identified from the different parts of As. triloba and As. parviflora samples. These compounds can be used to distinguish Asimina species. However, for definite identification of these unknown components, further investigation is required. This may provide a model for the rapid screening and structural characterization of bioactive constituents from plant extracts in a single analysis.

Differences in the chemical profiles and biological activities of Paeonia lactiflora and Paeonia obovata.

Paeonia lactiflora and P. obovata are perennial herbs, each root of which has been consumed as a major oriental medicine, Paeoniae Radix and a famous folk medicine, Mountain Paeony Root, respectively. Although morphological studies have been performed comparing these two plants, there is insufficient scientific evidence that characterizes the differences in their chemical profiles and biological activities. Hence, the present study was undertaken to compare these two medicinal foods using a high-performance liquid chromatography-diode-array detector (HPLC-DAD) analysis and a gastric ulcer model in mice. HPLC analysis employed to assess the nine components revealed that P. lactiflora exhibited higher contents of phenolic compounds than P. obovata. Although a monoterpene glycoside, 6'-O-acetylpaeoniflorin was identified in P. obovata, it was not detected in P. lactiflora. Multivariate statistical analysis for HPLC data revealed that the orthogonal projections to latent structure-discriminant analysis is more appropriate than principal component analysis for differentiating the two groups. Moreover, the 50% methanol P. lactiflora extract (PL) was more effective against experimental gastric ulcer than P. obovata extract (PO) in the HCl/ethanol-induced ulcer model. In addition, PL displayed higher 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging activity and lower nitric oxide production in a murine macrophage cell line, RAW 264.7, than PO. The DPPH radical scavenging activity of PL was as high as that of the positive control, butylated hydroxytoluene, at a concentration of 25 µg/mL.