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A rapid and sensitive bioassay based on liquid chromatography tandem mass spectrometry (LC-MS/MS) has been developed and validated for the simultaneous determination of eight coumarins in rat plasma. The liquid-liquid extraction method with ethyl acetate was used to prepare the plasma samples after addition of warfarin as an internal standard (IS). Chromatographic separation was performed on an Eclipse plus C18 column (100mm×4.6mm, 1.8μm) using gradient elution when 1mM ammonium acetate aqueous solution - acetonitrile was used as the mobile phase. The lower limit of quantitation (LLOQ) of each coumarin was lower than 2.16ngmL(-1). Intra-day and inter-day precisions were less than 15%. The accuracies were in the range of 88.9-117%. The mean recoveries of coumarins and IS were higher than 84%. The method was successfully applied to a pharmacokinetic study of eight coumarins in rats after oral administration of radix angelicae pubescentis., (Crown Copyright © 2013. Published by Elsevier B.V. All rights reserved.)

Objective: To establish a high-performance liquid chromatography (HPLC)-based method for determining the contents of psoralene, bergapten and apigenin in Ficus hirta Vahl., Methods: A Hypersil C18 column (250 mm × 4.6 mm, 5 µm) was used with the mobile phase of methanol-water (60:40), flow rate of 1.0 ml/min, detection wavelength of 268.7 nm and column temperature of 30 degrees celsius., Results: The calibration curve was linear within the range of 10.0-30.0 µg/ml for psoralene (r=0.9998), 15.0-45.0 µg/ml for bergapten (r=0.9998) and 5.0-15.0 µg/ml for apigenin (r=0.9992). The average recovery of psoralene was 99.7% (RSD=1.99%), that of bergapten was 99.9% (RSD=1.71%) and that of apigenin was 100.3% (RSD=1.78%)., Conclusion: The method is simple, economic and accurate with good reproducibility for the contents of psoralene, bergapten and apigenin.

A sensitive, specific and rapid liquid chromatography-mass spectrometry (LC-MS) method has been developed and validated for the simultaneous determination of xanthotoxin (8-methoxypsoralen), psoralen, isoimpinellin (5,8-dimethoxypsoralen) and bergapten (5-methoxypsoralen) in rat plasma using pimpinellin as an internal standard (IS). The plasma samples were pretreated by protein precipitation with methanol and chromatographic separation was performed on a C(18) column with a mobile phase composed of 1 mmol ammonium acetate and methanol (30:70, v/v). The detection was accomplished by multiple-reaction monitoring (MRM) scanning via electrospray ionization (ESI) source operating in the positive ionization mode. The optimized mass transition ion-pairs (m/z) for quantitation were 217.1/202.1 for xanthotoxin, 187.1/131.1 for psoralen, 247.1/217.0 for isoimpinellin, 217.1/202.1 for bergapten, and 247.1/231.1 for IS. The total run time was 6 min between injections. The calibration curves were linear over the investigated concentration range with all correlation coefficients higher than 0.998. The lower limits of quantitation (LLOQ) of these analytes were less than 1.21 ng/ml. The intra- and inter-day RSD were no more than 9.7% and the relative errors were within the range of -8.1% to 4.5%. The average extraction recoveries for all compounds were between 90.7% and 106.2%. The proposed method was further applied to the determination of actual plasma samples from rats after oral administration of Radix Glehniae extract., (Copyright 2010 Elsevier B.V. All rights reserved.)

Purpose: To discuss the contribution of psoralen and bergapten metabolites on psoralens toxicity., Methods: Computational chemistry prediction of metabolic reactions and toxicophoric groups based on the expert systems Derek and Meteor., Results: a total of 15 metabolites were suggested for both psoralen and bergapten based on phase 1 and 2 biotransformations until the 3rd generation. Five toxicophoric substructures were shared among psoralen, bergapten and their corresponding metabolites; one toxicophoric marker (resorcinol) was only identified in bergapten and its biotransformation products., Conclusion: Although the toxic effects of psoralens are well known and documented, there is little information concerning the role of their metabolites in this process. We believe this work add to the knowledge of which molecular substructures are relevant to the process of metabolism and toxicity induction, thus guiding the search and development of more effective and less toxic drugs to treat vitiligo.

A method for sample preparation and analysis by high-performance liquid chromatography with UV detection (HPLC-UV) was developed for analysis of psoralen, bergapten and 5-[3-(4,5-dihydro-5,5-dimethyl-4-oxo-2-furanyl)-butoxy]-7H-furo[3-2-g][1]benzopyran-7-one in capsules and tablets employed in Brazil for certain illnesses. The linearity, accuracy, the inter- and intra-day precision of the procedure were evaluated. Analytical curves for furanocoumarins were linear in the range of 1.0-50.0 microg/ml. The recoveries of the furanocoumarins in the products analyzed were 97.3-99.5%, and the percent coefficient of variation for the quantitative analysis of the furanocoumarins in the analyses was under 5%. For inter-equipment study gas chromatography (GC) was employed.

CYP6B1v1 is the principal cytochrome P450 monooxygenase (P450) that detoxifies dietary furanocoumarins in the guts of Papilio polyxenes, the black swallowtail caterpillar. Sequence alignments and structure comparisons of CYP6B1v1 with the mouse CYP2A5 and bacterial CYP102 proteins, which are also capable of metabolizing the linear furanocoumarin xanthotoxin (8-methoxypsoralen), suggested that Phe116, His117, Val368 and Phe484 might be active site residues. In a homology model developed for CYP6B1v1, the side chains of Phe116 and His117 located in the B'-C loop of SRS1 are predicted to be positioned above the haem plane, while the side chain of Phe484 located in SRS6 is predicted near the entrance of the catalytic pocket. Site-directed mutagenesis of residues Phe116, His117 and Phe484 indicated that these residues represent several of those that determine this protein's stability and substrate specificity. Whereas all aromatic mutants of Phe116 and Phe484 generated CO-difference spectra with maxima at 450 nm indicative of correctly configured monooxygenases, aromatic mutants of Phe116 exhibited reduced reactivities toward some furanocoumarins and aromatic mutants of Phe484 eliminated all reactivities toward furanocoumarins. All single and double aliphatic mutants of Phe116, His117 and Phe484 and aromatic mutants of His117 generated carbon monoxide (CO) difference spectra with maxima at 420 nm (P420) indicative of incorrectly configured monooxygenases. These studies define residues Phe116, His117 and Phe484 as determinants of this insect P450's catalytic site integrity and residues Phe116 and Phe484 as determinants of its substrate specificity. Conservation of Phe116 and His117 in an array of lepidopteran CYP6B proteins implies that these amino acids serve a similar function in other monooxygenases of the insect CYP6B subfamily.

A method for sample preparation and high-performance liquid chromatography detection and quantification has been developed for the routine analysis of psoralen and bergapten, photosensitizing compounds, in topic solutions employed in Brazil for treatment of vitiligo and psoriasis. The linearity, accuracy, the inter- and intra-day precision of the procedure were evaluated. The calibration curve for psoralen and bergapten were linear in the range 1.0-20.0 microg ml(-1). The best recoveries of the psoralens in the topic solutions analysed were 96.3-98.2%. The percentage coefficient of variation of the quantitative analysis of the psoralens in the product analyses was within 5%.

Dorstenin, 5-[3-(4,5-dihydro-5,5-dimethyl-4-oxo-2-furanyl)-butoxy]-7H-furo[3, 2-g] benzopyran-7-one, is a psoralen analog recently isolated from Dorstenia species (Moraceae). In order to characterize its biological activity, its photosensitizing and mutational properties were measured in wild-type E. coli and S. cerevisiae and also in strains carrying mutations which affect DNA repair. Compared to the high activities of psoralen and bergapten, dorstenin showed lower genotoxic effect.

The natural occurrence of biologically active furanocoumarins in common vegetables is an area of increasing interest with respect to human health. In this study, an efficient, rugged, and sensitive liquid chromatographic method with ultraviolet photodiode array detection was developed for the estimation of 5 biologically active furanocoumarins (psoralen, bergapten, xanthotoxin, trioxsalen, and angelicin) in celery and parsnips. When authentic samples were spiked with a mixture of furanocoumarins at individual levels of 2 to 10 microg/g, the method produced overall recoveries of 77 and 75% of all furanocoumarins from celery and parsnips, respectively. The method was applied in 2 laboratories to a multiyear survey of more than 200 samples. Of 110 parsnips samples, 109 (99%) contained quantitatable levels of furanocoumarins. The mean level of total furanocoumarins in the positive parsnip samples was 15.1 microg/g; the maximum level detected was 145 microg/g. Of 114 celery samples, 88 (77%) contained quantitatable levels of furanocoumarins. The mean level of total furanocoumarins in the positive celery samples was 1.9 microg/g; the maximum level detected was 15.2 microg/g. Xanthotoxin and bergapten were the most commonly detected furanocoumarins in both celery (68 and 63%) and parsnips (97 and 96%). Xanthotoxin had the highest mean level of positives in both celery (1.3 microg/g) and parsnips (8.5 microg/g). Little year-to-year variation in either total furanocoumarin levels or incidence was noted.

5,7-dimethoxycoumarin and isopimpinellin, together with the well-known phototoxic, photo-irritant furanocoumarins psoralen and 8-methoxypsoralen, were isolated and identified from leaves and fruits of Pelea anisata H. Mann, a plant whose fruit are used in the construction of mohikana leis used in parts of the Hawaiian Islands. The presence of phototoxic furanocoumarins would explain the occurrence of photodermatitis in the wearers of the leis.