Barbara Pendry

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.

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.

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 (</= 0.92 mg/mL). CONCLUSION These results have implications both for the manufacturers of commercial tinctures and also for herbal practitioners in the choice of tinctures for treating Herpes simplex infection.

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, Balds 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.