Croton stipulaceus Kunth, a native Mexican medicinal plant with antioxidant and anti-inflammatory activities / Croton stipulaceus Kunth, planta medicinal mexicana con actividad antioxidante y antiinflamatoria

Leaves of Croton stipulaceuswere extracted (EHex, ECHCl3and EEtOH extracts) to assesstheir antioxidant potential, anti-inflammatory activity in murine models and acute toxicity. EEtOH showed the highest effect in DPPH (37.80% inhibition), FRAP (1065.00 ± 55.30 µmolFe2+) and total polyphenols (231.24 ± 9.05 meq AG/gM). EHex was the most active, ~ 50% inhibition of TPA-induced ear edema; while EEtOH (dose of 2 mg/ear) showed the highest inhibition in the chronic model (97% inhibition), and inhibited MPO activity (48%). In carrageenan-induced edema, ECHCl3(dose 500 mg/kg) was the most active. None of the extracts showed acute toxicity (LD50) at 2 g/kg (p.o.). This work is the first report that supports the traditional use of C. stipulaceusas an anti-inflammatory.

De las hojas de Croton stipulaceusse obtuvieron diferentes extractos (EHex, ECHCl3y EEtOH) evaluando el potencial antioxidante y la actividad antiinflamatoria en modelos murinos y la toxicidad aguda. El EEtOH mostró mayor efecto en DPPH (37.80% inhibición), FRAP (1065.00 ± 55.30 µmolFe2+) y polifenolestotales (231.24 ± 9.05 meq AG/gM). El EHex fue el más activo, cercano al 50% de inhibición del edema auricular inducido con TPA; mientras que el EEtOH (dosis de 2 mg/oreja) mostró la mayor inhibición en el modelo crónico (97% inhibición), e inhibió la actividad de la MPO (48%). En el edema inducido con carragenina, el ECHCl3(dosis 500 mg/kg) fue el más activo. Ninguno de los extractos mostró una toxicidad aguda (DL50) mayor a 2 g/kg (p.o). Este trabajo es el primer reporte que sustenta el uso tradicional de C. stipulaceuscomo antiinflamatorio.

Sonchus oleraceus L.: ethnomedical, phytochemical and pharmacological aspects.

Sonchus oleraceus L. (Asteraceae) is a cosmopolitan species native to Europe commonly known as lettuce, sowthistle, chicory, or fake dandelion, considered a weed. However, for many years in various cultures around the world, it has been used as food and medicinal plant. The aim of this integrative review is to document the ethnomedical, phytochemical, and pharmacological information of this species. Forty-one papers document the use of S. oleraceus to heal of a wide variety of diseases. However, gastrointestinal problems, diabetes, inflammation, infections, hepatitis, wounds, and to consume it as food are the most common uses. On the other hand, only 11 items highlight that the main groups of secondary metabolites in this species are flavonoids and terpene lactones. Finally, 45 items reveal that antioxidant, antimicrobial, antiproliferative and cytotoxic were the most studied pharmacological activities. In vitro and in vivo studies of extracts and components isolated from different parts of S. oleraceus have provided a concrete overview of the pharmacological properties of this species that supports its ethnomedical uses in cultures from different parts of the world. The reports of this species have focused solely on the study of the complete plant, leaves, and aerial parts, so it is necessary to study other parts of this species to search for bioactive compounds. No clinical studies were found, which creates an opportunity to expand scientific knowledge of this species.

Flaxseed Oil (Linum usitatissimum) Prevents Cognitive and Motor Damage in Rats with Hyperammonemia.

Hyperammonemia is characterized by the excessive accumulation of ammonia in the body as a result of the loss of liver detoxification, leading to the development of hepatic encephalopathy (HE). These metabolic alterations carry cognitive and motor deficits and cause neuronal damage, with no effective treatment at present. In this study, we aimed to evaluate the effect of two subacute oral administrations of flaxseed oil (0.26 and 0.52 mL/kg) on short- and long-term memory, visuospatial memory, locomotor activity, motor coordination, and the neuronal morphology of the prefrontal cortex (PFC) via tests on Wistar rats with hyperammonemia. The goal was to identify its role in the regulation of cerebral edema, without liver damage causing cerebral failure. In contrast with an ammonium-rich diet, flaxseed oil and normal foods did not cause cognitive impairment or motor alterations, as evidenced in the short-term and visuospatial memory tests. Furthermore, the flaxseed oil treatment maintained a regular neuronal morphology of the prefrontal cortex, which represents a neuroprotective effect. We conclude that the oral administration of flaxseed oil prevents cognitive and motor impairments as well as neuronal alterations in rats with hyperammonemia, which supports the potential use of this oil to ameliorate the changes that occur in hepatic encephalopathy.

C-Phycocyanin: A Phycobiliprotein from Spirulina with Metabolic Syndrome and Oxidative Stress Effects.

Spirulina maxima is a cyanobacterium considered a "superfood" due to its metabolites and nutrient content. These include a complex mixture of minerals, vitamins, fatty acids, proteins, and accessory pigments. In recent years, it has positioned itself as a promising source of bioactive molecules for the treatment of several diseases, including metabolic syndrome, coronary diseases, cancer, and the improvement of health modulating oxidative stress. C-Phycocyanin (C-PC) is a photosynthetic pigment from green-blue cyanobacterium and the most abundant phycobiliprotein in the Spirulina genus with various pharmacological properties attributed due to its antioxidant capacity but has no specific cellular target. This has made it a molecule of great interest in biomedical research. This review focuses on the pharmacological effects and the benefits on metabolic syndrome and oxidative stress of C-PC.

Consideraciones bioéticas para la investigación científica de plantas medicinales contra el cáncer en México / Bioethical considerations for scientific research on medicinal plants against cancer in Mexico

El presente artículo de reflexión revisa aspectos bioéticos en estudios científicos de plantas utilizadas en medicina tradicional para el tratamiento del cáncer en México. El conocimiento de las plantas medicinales documentadas en la medicina tradicional se relaciona con enfermedades como el cáncer. Esta enfermedad presenta altas tasas de mortalidad en el mundo, lo que incrementa la necesidad de nuevos fármacos para la quimioterapia. Aquí las plantas medicinales juegan un papel importante. Así, al ejecutar un estudio de plantas medicinales se deben considerar aspectos bioéticos fundamentales para la medicina tradicional como el muestreo, el estudio fitoquímico y biológico en líneas celulares de cáncer, basándose en la normatividad vigente, en estándares internacionales del Instituto Nacional del Cáncer y, principalmente, en los cuatro principios básicos de la bioética. Concluimos que es tarea de la comunidad científica avalar o desmentir el uso tradicional de las plantas medicinales para el tratamiento de enfermedades. El estudio de especies vegetales debe cumplir criterios donde la teoría, la investigación y la práctica ofrezcan soluciones a largo plazo y la protección ética de estas, del conocimiento tradicional y de los pacientes que recurren a esta alternativa.

The present reflection paper reviews bioethical aspects in scientific studies of plants used in traditional medicine for the treatment of cancer in Mexico. The knowledge of medicinal plants documented in traditional medicine is related to diseases such as cancer. This disease has high mortality rates worldwide, increasing the need for new chemotherapy drugs. Here medicinal plants play an important role. Thus, when executing a study of medicinal plants, fundamental bioethical aspects for traditional medicine such as sampling, phytochemical and biological study in cancer cell lines, based on current regulations, international standards of the National Cancer Institute and, mainly, the four basic principles of bioethics, should be considered. We conclude that it is the task of the scientific community to endorse or deny the traditional use of medicinal plants for the treatment of diseases. The study of plant species must meet criteria where theory, research and practice offer long-term solutions and the ethical protection of plant species, traditional knowledge and patients who resort to this alternative.

Chemometrics: a complementary tool to guide the isolation of pharmacologically active natural products.

Chemometrics offers an important complementary tool to enhance the searching and isolation of bioactive natural products from natural sources.

Antioxidant, anti-inflammatory and antinociceptive potential of Ternstroemia sylvatica Schltdl. & Cham.

OBJECTIVE: To evaluate the anti-inflammatory, analgesic, antioxidant and acute toxicity of extracts obtained from a successive extraction with solvents of ascending polarity [hexane, hex; chloroform, CHCl3 and ethanol (EtOH)] of Ternstroemia sylvatica Schltdl. & Cham. METHODS: The antioxidant potential was evaluated by 2,2 diphenyl-1-picrylhydrazyl, the ferric reducing/antioxidant power assays and by determining the total phenolic content. The anti-inflammatory and antinociceptive effects were evaluated using the in vivo croton oil-induced ear edema, phorbol 12-myristate 13-acetate induced ear edema, carrageenan-induced paw edema, acetic acid-induced writhing and formalin murine models. The acute toxicity was tested using the Lorke's method in mice. RESULTS: The EtOH extract was the most active for the antioxidant potential tests diphenyl-1-picrylhydrazyl (68.70% inhibition), ferric reducing/antioxidant power [(2431.30 ± 102.10) mmol Fe2+ and total polyphenols content (215.80 ± 8.50) meqAG/g]. The anti-inflammatory activity was evaluated by topical application of croton oil (2 mg/ear dose) where the EtOH extract showed the strongest activity compared to the control group (45.13% inhibition), whereas in the phorbol 12-myristate 13-acetate model, at the same dose, the CHCl3 extract showed the highest inhibition (42.88%). In the carrageenan induced edema model, the EtOH extract showed a stronger inhibition compared to indomethacin (56.34% and 50.70% at doses of 250 and 500 mg/kg of extract, respectively) during the first hour. Similarly, the same extract showed the highest analgesic activity (30.60% inhibition) in the acetic acid contortion assay, and in the formalin test it showed a greater effect with respect to the control group in both phases. CONCLUSIONS: Our work confirms the value of Ternstroemia sylvatica as an important anti-inflammatory and analgesic plant, whose mechanism seems to be associated to its antioxidant effects, and supports its uses in the Mexican traditional medicine.

Chemometrics-enhanced high performance liquid chromatography-ultraviolet detection of bioactive metabolites from phytochemically unknown plants.

This work describes the use of Colubrina greggii as a model to investigate the use of chemometric analysis combined with data from a leishmanicidal bioassay, using Principal Component Analysis (PCA) and Orthogonal Projections to Latent Structures (O-PLS), to detect biologically active natural products in crude extracts from plants having little or no phytochemical information. A first analysis of the HPLC-UV profiles of the extract and its semi-purified fractions using both Principal Component Analysis (PCA) and Orthogonal Partial Least Squares (O-PLS) indicated that the components at tR 48.2, 48.7, 51.8min correlated with the variation in bioactivity. However, a further O-PLS analysis of the HPLC-UV profiles of fractions obtained through a final semi-preparative HPLC purification showed two components at tR 48.7 and 49.5min which correlated with the variation of the bioactivity in a high performance predictive model, with high determination coefficient, high correlation coefficient values (R(2) and Q(2)=0.99) and a low root mean square error (RMSE=0.018). This study demonstrates that the association of chemometric analysis with bioassay results can be an excellent strategy for the detection and isolation of bioactive metabolites from phytochemically unknown plant crude extracts.

Salvinorin A content in legal high products of Salvia divinorum sold in Mexico.

Salvia divinorum (Lamiaceae) is a herb native to Mexico where it is used by Mazatec shamans for spiritual and divination purposes. S. divinorum products are easily available to consumers and are used worldwide as legal highs because of the hallucinogenic effects caused mainly by salvinorin A. Highly popular videos and websites on the internet depicting the use of S. divinorum products have contributed to an increase in their consumption. Recent reports have highlighted the potential of these products to induce psychosis in consumers. In Mexico, dried leaf extracts of S. divinorum are sold in different strengths, claiming to correlate with increasing amounts of salvinorin A. In order to determine the variability of salvinorin A content between brands and to investigate possible correlation between brand strengths, this study sought to quantify salvinorin A in commercial products available in Mexico using an HPLC method. The HPLC analytical method showed a correlation coefficient R(2)>0.99, with LOD of 0.44 µg/mL and LOQ of 1.34 µg/mL. The retention time for salvinorin A was 23.09±0.95 min and the measured concentrations ranged between 8.32±0.65 and 56.52±3.77 mg/g dried leaf. The results for brand c did not show an agreement between the declared and the calculated amount of salvinorin A. Additionally, the emergence in Mexico of high strength salvia products (100×), the lack of regulation and the observed variability of salvinorin A content between brands of commercial legal highs products of S. divinorum could result in a health problem for consumers.

Cnidoscolus chayamansa Mc Vaugh, an important antioxidant, anti-inflammatory and cardioprotective plant used in Mexico.

ETHNOPHARMACOLOGICAL RELEVANCE: Cnidoscolus chayamansa Mc Vaugh (Euphorbiaceae) is commonly known as 'chaya' in Central America. In South East Mexico, because of its high nutritional values, is an important part of the diet of many indigenous communities. Chaya is also used as a traditional remedy for the treatment of diabetes, rheumatism, gastrointestinal disorders and inflammation-related diseases. Although Cnidoscolus chayamansa is one of most used and valued medicinal plants, only few studies on documenting its pharmacological properties can be found. MATERIALS AND METHODS: Dried leaves of Cnidoscolus chayamansa were subjected to a successive maceration using Hex, EtOAc and EtOH. The antioxidant activities of the extracts were tested using the DPPH radical scavenging, Ferric reducing/antioxidant power and total phenolic content assays. To determine the anti-inflammatory activity, the TPA-induced mouse ear edema and the carrageenan-induced mouse paw edema assays were used. The cardioprotective effects of the EtOH extract was determined using the ischemia/reperfusion (I/R) rat model. Finally, the acute toxicity was determined using Lorke's method. RESULTS: The results showed a similar anti-inflammatory activity (≈30%) for all extracts but only the EtOAc extract showed relevant activity when applied intraperitoneally. When tested for their antioxidant activity none of the extracts showed a significant activity suggesting that the antinflammatory activity is not related to a direct free radical scavenging of the extracts. Additionally, the EtOH extract showed a strong cardioprotective effect at 500mg/kg when given orally. Both the EtOAc and the EtOH extract have a LD50 >5g/kg, confirming their safety in acute oral administration. CONCLUSIONS: All these results are relevant for a better understanding of the therapeutic used of Cnidoscolus chayamansa in the Mexican traditional medicine and highlights its cardioprotective potential.

Antimicrobial assays of three native British plants used in Anglo-Saxon medicine for wound healing formulations in 10th century England.

ETHNOPHARMACOLOGICAL RELEVANCE: Three important Anglo-Saxon medical texts from the 10th century contain herbal formulations for over 250 plant species, many of which have yet to be evaluated for their phytochemical and/or pharmacological properties. In this study, three native British plants were selected to determine antimicrobial activity relevant to treating bacterial infections and wounds. MATERIALS AND METHODS: Several preparations of Agrimonia eupatoria L., Arctium minus (Hill) Bernh. and Potentilla reptans L. were screened for antimicrobial activity against selected Gram-positive and Gram-negative bacteria of relevance in wounds using a 96 well plate microdilution method (200, 40 and 8µg/mL). Minimum inhibitory concentration (MIC) values were determined for the most potent extracts from 2 to 0.004mg/mL and HPLC chromatograms examined by multivariate analysis. Principle components analysis (PCA) was used to identify chemical differences between antimicrobial activity of the crude extracts. RESULTS: The HPLC-PCA score plots attributed HPLC peaks to the antimicrobial activity with all three plants inhibiting growth of Gram-positive Staphylococcus aureus by >50% in four or more extracts. The first two principal components (PC) represented 87% of the dataset variance. The P. reptans 75% ethanol root extract exhibited the greatest range of activity with MIC(50) at 31.25µg/mL to a total MIC that was also the minimum bactericidal concentration (MBC) at 1mg/mL. Additionally, the root of P. reptans, inhibited growth of Gram-negative bacteria with the 75% ethanol extract having a MIC(50) at 1mg/mL against Pseudomonas aeruginosa and the decoction a MIC(50) at 3.9µg/mL against Escherichia coli. CONCLUSIONS: The results indicate a moderate antimicrobial activity against common wound pathogens for P. reptans suggesting it may well have been effective for treating wound and bacterial infections. Anglo-Saxon literary heritage may provide a credible basis for researching new antimicrobial formulations. Our approach encompassing advanced analytical technologies and chemometric models paves the way for systematic investigation of Anglo-Saxon medical literature for further therapeutic indications to uncover knowledge of native British plants, some of which are currently lost to modern Western herbal medicine.

Anglo-Saxon pharmacopoeia revisited: a potential treasure in drug discovery.

Three of the four major Anglo-Saxon collections reporting medicinal formulations in England from the 10th century, the Old English Herbarium, Bald's Leechbook and the Lacnunga, could contain leads and insights into new medicinal uses. Previous pharmacological studies of medicinal plants mentioned in Anglo-Saxon medical texts suggested that some were effective and led to the identification and isolation of natural compounds. For example, matricin from yarrow Achillea millefolium L., is a proprionic acid analogue that yields chamazulene carboxylic acid with cyclooxygenase-2 activity similar to that of ibuprofen. As we discuss here, multidisciplinary projects could further explore historical texts to discover additional plant metabolites with potential pharmacological applications.

The ethanol extract of Scutellaria baicalensis and the active compounds induce cell cycle arrest and apoptosis including upregulation of p53 and Bax in human lung cancer cells.

Despite a lack of scientific authentication, Scutellaria baicalensis is clinically used in Chinese medicine as a traditional adjuvant to chemotherapy of lung cancer. In this study, cytotoxicity assays demonstrated that crude ethanolic extracts of S. baicalensis were selectively toxic to human lung cancer cell lines A549, SK-LU-1 and SK-MES-1 compared with normal human lung fibroblasts. The active compounds baicalin, baicalein and wogonin did not exhibit such selectivity. Following exposure to the crude extracts, cellular protein expression in the cancer cell lines was assessed using 2D gel electrophoresis coupled with MALDI-TOF-MS/Protein Fingerprinting. The altered protein expression indicated that cell growth arrest and apoptosis were potential mechanisms of cytotoxicity. These observations were supported by PI staining cell cycle analysis using flow cytometry and Annexin-V apoptotic analysis by fluorescence microscopy of cancer cells treated with the crude extract and pure active compounds. Moreover, specific immunoblotting identification showed the decreased expression of cyclin A results in the S phase arrest of A549 whereas the G(0)/G(1) phase arrest in SK-MES-1 cells results from the decreased expression of cyclin D1. Following treatment, increased expression in the cancer cells of key proteins related to the enhancement of apoptosis was observed for p53 and Bax. These results provide further insight into the molecular mechanisms underlying the clinical use of this herb as an adjuvant to lung cancer therapy.

Identification of sakurasosaponin as a cytotoxic principle from Jacquinia flammea.

The crude ethanolic extract of leaves, stem-bark and roots of J. flammea were tested for their cytotoxic effect against two mammalian cell lines (HeLa and RAW 264.7) and four bacterial species (Staphylococcus aureus, Bacillus subtilis, Escherichia coli and Pseudomonas aeruginosa). When tested at the concentration of 100 microg/mL, the root extract showed the highest cytotoxic activity against mammalian cells followed by the stem-bark extract while the leaves extract did not show significant activity. No antibacterial activity was detected for all extracts when tested up to 500 microg/disc in the disc diffusion assay. The cytotoxic root extract was subjected to fractionation using solvents of ascending polarity: petroleum ether, chloroform, ethylacetate, butanol and water. The water fraction which showed cytotoxic activity was further subjected to routine bioassay-guided fraction to lead to the isolation of sakurasosaponin as the active principle. The recorded IC50 value for sakurasosaponin was 11.3 +/- 1.52 and 3.8 +/- 0.25 microM (n=3) against HeLa and RAW 264.7 respectively. The identification of sakurasosaponin was based on analysis of spectroscopic data.

Triterpenoid saponins from a cytotoxic root extract of Sideroxylon foetidissimum subsp. gaumeri.

Evaluation of the cytotoxicity of an ethanolic root extract of Sideroxylonfoetidissimum subsp. gaumeri (Sapotaceae) revealed activity against the murine macrophage-like cell line RAW 264.7. Systematic bioassay-guided fractionation of this extract gave an active saponin-containing fraction from which four saponins were isolated. Use of 1D ((1)H, (13)C, DEPT135) and 2D (COSY, TOCSY, HSQC, and HMBC) NMR, mass spectrometry and sugar analysis gave their structures as 3-O-(beta-D-glucopyranosyl-(1-->6)-beta-D-glucopyranosyl)-28-O-(alpha-L-rhamnopyranosyl-(1-->3)[beta-D-xylopyranosyl-(1-->4)]-beta-D-xylopyranosyl-(1-->4)-alpha-L-rhamnopyranosyl-(1-->2)-alpha-L-arabinopyranosyl)-16alpha-hydroxyprotobassic acid, 3-O-beta-D-glucopyranosyl-28-O-(alpha-L-rhamnopyranosyl-(1-->3)[beta-D-xylopyranosyl-(1-->4)]-beta-D-xylopyranosyl-(1-->4)-alpha-L-rhamnopyranosyl-(1-->2)-alpha-L-arabinopyranosyl)-16alpha-hydroxyprotobassic acid, 3-O-(beta-D-glucopyranosyl-(1-->6)-beta-D-glucopyranosyl)-28-O-(alpha-L-rhamnopyranosyl-(1-->3)-beta-D-xylopyranosyl-(1-->4)[beta-D-apiofuranosyl-(1-->3)]-alpha-L-rhamnopyranosyl-(1-->2)-alpha-L-arabinopyranosyl)-16alpha-hydroxyprotobassic acid, and the known compound, 3-O-beta-D-glucopyranosyl-28-O-(alpha-L-rhamnopyranosyl-(1-->3)[beta-D-xylopyranosyl-(1-->4)]-beta-D-xylopyranosyl-(1-->4)-alpha-L-rhamnopyranosyl-(1-->2)-alpha-L-arabinopyranosyl)-protobassic acid. Two further saponins were obtained from the same fraction, but as a 5:4 mixture comprising 3-O-(beta-D-glucopyranosyl)-28-O-(alpha-L-rhamnopyranosyl-(1-->3)-beta-D-xylopyranosyl-(1-->4)[beta-D-apiofuranosyl-(1-->3)]-alpha-L-rhamnopyranosyl-(1-->2)-alpha-L-arabinopyranosyl)-16alpha-hydroxyprotobassic acid and 3-O-(beta-D-apiofuranosyl-(1-->3)-beta-D-glucopyranosyl)-28-O-(alpha-L-rhamnopyranosyl-(1-->3)[beta-D-xylopyranosyl-(1-->4)]-beta-D-xylopyranosyl-(1-->4)-alpha-L-rhamnopyranosyl-(1-->2)-alpha-L-arabinopyranosyl)-16alpha-hydroxyprotobassic acid, respectively. This showed greater cytotoxicity (IC(50)=11.9+/-1.5 microg/ml) towards RAW 264.7 cells than the original extract (IC(50)=39.5+/-4.1 microg/ml), and the saponin-containing fraction derived from it (IC(50)=33.7+/-6.2 microg/ml).

Validation of a HPLC method for flavonoid biomarkers in skullcap (Scutellaria) and its use to illustrate wide variability in the quality of commercial tinctures.

PURPOSE: To compare the flavonoid biomarker content (baicalin, baicalein and wogonin) of eleven commercial tinctures derived from Scutellaria lateriflora aerial parts (n=7) and Scutellaria baicalensis root (n=4). S. lateriflora tinctures are used in by western herbal practitioners to treat anxiety whereas S. baicalensis tinctures are used to treat inflammatory disease. METHODS: Baicalin and baicalein were purchased from Aldrich Chemical Co. and Wogonin was purchased from ChromaDex. The internal standard (4-hydroxybenzoic acid) was obtained from Acros Organics. The column used was a Luna C18, 5 m (150 x 4.6 mm, Phenomenex) maintained at ambient room temperature. A HP1050 HPLC system was used, comprising a gradient pump with degasser, a variable wavelength UV detector set to 270 nm, and an autosampler. Gradient elution was performed using 0.1% formic acid (eluent A) and methanol (eluent B). The gradient elution initial conditions were 45% B with linear gradient to 60% from 2 to 10 min, followed by linear gradient to 70% B at 30 min, and then linear gradient to 99% B at 31 min, this proportion being maintained for 1 min. The mobile phase was then returned to initial conditions at 33 min and maintained until the end of the run at 35 min. The flow rate was 1 mL/min. The assay was validated for sensitivity, accuracy and reproducibility. RESULTS: The concentration range of biomarkers (baicalin, baicalein and wogonin) in commercial tinctures is reported for S. lateriflora (baicalin: 0-12.66 mg/mL; baicalein: 0-0.63 mg/mL; wogonin: 0-0.16 mg/mL) and for S. baicalensis (baicalin: 0.12-10.61 mg/mL; baicalein: 0.52-5.88 mg/mL; wogonin: 0.08-1.61 mg/mL). CONCLUSION: The wide variability in biomarker concentrations between commercial tinctures has important implications for the manufacturers of commercial tinctures, for herbal practitioners in the choice of tinctures and not least for pharmacology and clinical researchers.

Comparison of rosmarinic acid content in commercial tinctures produced from fresh and dried lemon balm (Melissa officinalis).

PURPOSE: To measure the rosmarinic acid content of eight commercial tinctures derived from fresh (n= 5) and dried (n=3) Melissa officinalis herb. METHODS: Rosmarinic acid and the internal standard (esculin) were purchased from Aldrich Chemical Co. The column used was a Luna C18, 5 um (150 x 4.6 mm I.D., Phenomenex) maintained at ambient room temperature. The HPLC system consisted of a Shimadzu SCL-6B controller, Shimadzu LC-6A pumps, Shimadzu SPD-6A UV single wavelength spectrophotometric detector set to 320 nm and Shimadzu SIL-6B autosampler. Gradient elution of the samples and standard were performed using ammonium formate (0.02 M; pH 6.25 at 27 oC; eluent A) and methanol (eluent B). The gradient elution initial conditions were 2% of eluent B with linear gradient to 60% at 30 min, followed by linear gradient to 90% of eluent B at 31 min, this proportion being maintained for 4 min. The column was then returned to the initial condition at 36 min and maintained until the end of the run at 43 min. The flow rate was 1 mL/min. The assay was validated for sensitivity, accuracy and reproducibility. RESULTS: The content of rosmarinic acid in commercial tinctures was significantly higher in the tinctures made from dried plant material (2.96 - 22.18 mg/mL) compared to fresh plant tinctures (