Joe-Ann McCoy

Hypericum in infection: Identification of anti-viral and anti-inflammatory constituents.

The Iowa Center for Research on Botanical Dietary Supplements seeks to optimize Echinacea, Hypericum, and Prunella botanical supplements for human-health benefit, emphasizing anti-viral, anti-inflammatory, and anti-pain activities. This mini-review reports on ongoing studies on Hypericum. The Center uses the genetically diverse, well-documented Hypericum populations collected and maintained at the USDA-ARS North Central Regional Plant Introduction Station (NCRPIS), and the strength of research in synthetic chemistry at Iowa State University to tap natural diversity, to help discover key constituents and interactions among constituents that impact bioactivity and toxicity. The NCRPIS has acquired more than 180 distinct populations of Hypericum, with a focus on Hypericum perforatum L. (Hypericaceae), representing about 13% of currently recognized taxa. Center chemists have developed novel synthetic pathways for key flavones, acyl phloroglucinols, hyperolactones, and a tetralin that have been found in Hypericum, and these compounds are used as standards and for bioactivity studies. Both light-dependent and light-independent anti-viral activities have been identified by using bioactivity-guided fractionation of H. perforatum and a HIV-1 infection test system. Our Center has focused on light-independent activity, potentially due to novel chemicals, and polar fractions are undergoing further fractionation. Anti-inflammatory activity has been found to be light-independent, and fractionation of a flavonoid-rich extract revealed four compounds (amentoflavone, chlorogenic acid, pseudohypericin, and quercetin) that interacted in the light to inhibit lipopolysaccharide-induced prostaglandin E2 activity. The Center continues to explore novel populations of H. perforatum and related species to identify constituents and interactions of constituents that contribute to potential health benefits related to infection.

Permeability of rosmarinic acid in Prunella vulgaris and ursolic acid in Salvia officinalis extracts across Caco-2 cell monolayers

ETHNOPHARMACOLOGICAL RELEVANCE Rosmarinic acid (RA), a caffeic acid-related compound found in high concentrations in Prunella vulgaris (self-heal), and ursolic acid (UA), a pentacyclic triterpene acid concentrated in Salvia officinalis (sage), have been traditionally used to treat inflammation in the mouth, and may also be beneficial for gastrointestinal health in general. AIM OF THE STUDY To investigate the permeabilities of RA and UA as pure compounds and in Prunella vulgaris and Salvia officinalis ethanol extracts across human intestinal epithelial Caco-2 cell monolayers. MATERIALS AND METHODS The permeabilities and phase II biotransformation of RA and UA as pure compounds and in herbal extracts were compared using Caco-2 cells with HPLC detection. RESULTS The apparent permeability coefficient (P(app)) for RA and RA in Prunella vulgaris extracts was 0.2 ± 0.05 × 10(-6)cm/s, significantly increased to 0.9 ± 0.2 × 10(-6)cm/s after β-glucuronidase/sulfatase treatment. P(app) for UA and UA in Salvia officinalis extract was 2.7 ± 0.3 × 10(-6)cm/s and 2.3 ± 0.5 × 10(-6)cm/s before and after β-glucuronidase/sulfatase treatment, respectively. Neither compound was affected in permeability by the herbal extract matrix. CONCLUSION RA and UA in herbal extracts had similar uptake as that found using the pure compounds, which may simplify the prediction of compound efficacy, but the apparent lack of intestinal glucuronidation/sulfation of UA is likely to further enhance the bioavailability of that compound compared with RA.

Inhibition of HIV-1 infection by aqueous extracts of Prunella vulgaris L.

BackgroundThe mint family (Lamiaceae) produces a wide variety of constituents with medicinal properties. Several family members have been reported to have antiviral activity, including lemon balm (Melissa officinalis L.), sage (Salvia spp.), peppermint (Mentha × piperita L.), hyssop (Hyssopus officinalis L.), basil (Ocimum spp.) and self-heal (Prunell a vulgaris L.). To further characterize the anti-lentiviral activities of Prunella vulgaris, water and ethanol extracts were tested for their ability to inhibit HIV-1 infection.ResultsAqueous extracts contained more anti-viral activity than did ethanol extracts, displaying potent antiviral activity against HIV-1 at sub μg/mL concentrations with little to no cellular cytotoxicity at concentrations more than 100-fold higher. Time-of-addition studies demonstrated that aqueous extracts were effective when added during the first five hours following initiation of infection, suggesting that the botanical constituents were targeting entry events. Further analysis revealed that extracts inhibited both virus/cell interactions and post-binding events. While only 40% inhibition was maximally achieved in our virus/cell interaction studies, extract effectively blocked post-binding events at concentrations similar to those that blocked infection, suggesting that it was targeting of these latter steps that was most important for mediating inhibition of virus infectivity.ConclusionsWe demonstrate that aqueous P. vulgaris extracts inhibited HIV-1 infectivity. Our studies suggest that inhibition occurs primarily by interference of early, post-virion binding events. The ability of aqueous extracts to inhibit early events within the HIV life cycle suggests that these extracts, or purified constituents responsible for the antiviral activity, are promising microbicides and/or antivirals against HIV-1.

Inhibition of lentivirus replication by aqueous extracts of Prunella vulgaris

BackgroundVarious members of the mint family have been used historically in Chinese and Native American medicine. Many of these same family members, including Prunella vulgaris, have been reported to have anti-viral activities. To further characterize the anti-lentiviral activities of P. vulgaris, water and ethanol extractions were tested for their ability to inhibit equine infectious anemia virus (EIAV) replication.ResultsAqueous extracts contained more anti-viral activity than did ethanol extracts, displaying potent anti-lentiviral activity against virus in cell lines as well as in primary cell cultures with little to no cellular cytotoxicity. Time-of-addition studies demonstrated that the extracts were effective when added during the first four h of the viral life cycle, suggesting that the botanical constituents were targeting the virion itself or early entry events. Further analysis revealed that the extracts did not destroy EIAV virion integrity, but prevented viral particles from binding to the surface of permissive cells. Modest levels of anti-EIAV activity were also detected when the cells were treated with the extracts prior to infection, indicating that anti-EIAV botanical constituents could interact with both viral particles and permissive cells to interfere with infectivity. Size fractionation of the extract demonstrated that eight of the nine fractions generated from aqueous extracts displayed anti-viral activity. Separation of ethanol soluble and insoluble compounds in the eight active fractions revealed that ethanol-soluble constituents were responsible for the anti-viral activity in one fraction whereas ethanol-insoluble constituents were important for the anti-viral activity in two of the other fractions. In three of the five fractions that lost activity upon sub-fractionation, anti-viral activity was restored upon reconstitution of the fractions, indicating that synergistic anti-viral activity is present in several of the fractions.ConclusionOur findings indicate that multiple Prunella constituents have profound anti-viral activity against EIAV, providing additional evidence of the broad anti-viral abilities of these extracts. The ability of the aqueous extracts to prevent entry of viral particles into permissive cells suggests that these extracts may function as promising microbicides against lentiviruses.

Comparison of Flow Injection MS, NMR, and DNA Sequencing: Methods for Identification and Authentication of Black Cohosh (Actaea racemosa)

Flow injection mass spectrometry and proton nuclear magnetic resonance spectrometry, two metabolic fingerprinting methods, and DNA sequencing were used to identify and authenticate Actaea species. Initially, samples of Actaea racemosa from a single source were distinguished from other Actaea species based on principal component analysis and soft independent modeling of class analogies of flow injection mass spectrometry and proton nuclear magnetic resonance spectrometry metabolic fingerprints. The chemometric results for flow injection mass spectrometry and proton nuclear magnetic resonance spectrometry agreed well and showed similar agreement throughout the study. DNA sequencing using DNA sequence data from two independent gene regions confirmed the metabolic fingerprinting results. Differences were observed between A. racemosa samples from four different sources, although the variance within species was still significantly less than the variance between species. A model based on the combined A. racemosa samples from the four sources consistently permitted distinction between species. Additionally, the combined A. racemosa samples were distinguishable from commercial root samples and from commercial supplements in tablet, capsule, or liquid form. DNA sequencing verified the lack of authenticity of the commercial roots but was unsuccessful in characterizing many of the supplements due to the lack of available DNA.

ECHINACEA SANGUINEA AND ECHINACEA PALLIDA EXTRACTS STIMULATE GLUCURONIDATION AND BASOLATERAL TRANSFER OF BAUER ALKAMIDES 8 AND 10 AND KETONE 24 AND INHIBIT P-GLYCOPROTEIN TRANSPORTER IN CACO-2 CELLS

The use of Echinacea as a medicinal herb is prominent in the United States, and many studies have assessed the effectiveness of Echinacea as an immunomodulator. We hypothesized that Bauer alkamides 8, 10, and 11 and ketone 24 were absorbed similarly either as pure compounds or from Echinacea sanguinea and Echinacea pallida ethanol extracts, and that these Echinacea extracts could inhibit the P-glycoprotein transporter in Caco-2 human intestinal epithelial cells. Using HPLC analysis, the permeation rate of Bauer alkamides by passive diffusion across Caco-2 cells corresponded with compound hydrophilicity (alkamide 8 > 10 > 11), independent of the plant extract matrix. Both Echinacea ethanol extracts stimulated apparent glucuronidation and basolateral efflux of glucuronides of alkamides 8 and 10 but not alkamide 11. Bauer ketone 24 was totally metabolized to more hydrophilic metabolites when administered as a single compound, but was also glucuronidated when present in Echinacea extracts. Bauer alkamides 8, 10, and 11 (175-230 µM) and ethanol extracts of E. sanguinea (1 mg/mL, containing ~ 90 µM total alkamides) and E. pallida (5 mg/mL, containing 285 µM total alkamides) decreased the efflux of the P-glycoprotein transporter probe calcein-AM from Caco-2 cells. These results suggest that other constituents in these Echinacea extracts facilitated the metabolism and efflux of alkamides and ketones, which might improve therapeutic benefits. Alkamides and Echinacea extracts might be useful in potentiating some chemotherapeutics, which are substrates for the P-glycoprotein transporter.