Chemical comparison and discrimination of two plant sources of Angelicae dahuricae Radix, Angelica dahurica and Angelica dahurica var. formosana, by HPLC-Q/TOF-MS and quantitative analysis of multiple components by a single marker.

INTRODUCTION: Angelica dahurica(BZ) and Angelica dahurica var. formosana(HBZ) are two plant sources of Angelicae dahuricae Radix. Although BZ and HBZ are commonly used herbal medicines with great medicinal and dietary values, study on their phytochemicals and bioactive compositions is limited. OBJECTIVE: To compare the chemical compositions of BZ and HBZ and find the chemical makers for discrimination and quality evaluation of the two botanical origins of Angelicae dahuricae Radix. METHODOLOGY: A high-performance liquid chromatography coupled with quadrupole time-of-flight mass spectrometry method was established for chemical profiling of BZ and HBZ. Then, a quantitative analysis of multiple components by a single marker method was developed for simultaneous determination of nine bioactive coumarins (xanthotoxol, oxypeucedanin hydrate, byakangelicin, xanthotoxin, bergapten, oxypeucedanin, phellopterin, imperatorin and isoimperatorin). Moreover, chemometrics were performed to compare and discriminate BZ and HBZ samples. RESULTS: A total of 30 coumarins compounds were identified, and the chemical compositions in BZ and HBZ were quite similar. The quantitative analysis showed that there were significant differences in the contents of bioactive coumarins, and the chemometric analysis indicated five coumarins (xanthotoxol, xanthotoxin, bergapten, phellopterin and isoimperatorin) were responsible for the significant differences between BZ and HBZ, which could be used as chemical markers to distinguish the two original plant sources of Angelicae dahuricae Radix. CONCLUSION: The present work provided useful information for understanding the chemical differences between BZ and HBZ and also provided feasible methods for quality evaluation and discrimination of herbal medicines originating from multiple botanical sources.

Investigations on the constituents of SagaPro tablets, a food supplement manufactured from Angelica archangelica leaf.

Saga Pro is a food supplement product manufactured in Iceland and marketed internationally. It is claimed to have anti-nocturia effect and the flavonoid isoquercitrin has been suggested to play a role in this assumed activity. The purpose of this study was to identify and quantify the main flavonoids and furanocoumarins in the SagaPro tablets and to evaluate the importance of their presence. Isoquercitrin was identified as a constituent in an amount of 158 µg/tablet. This is a p.o. dosage highly unlikely to have an effect on nocturia or any other pharmacologically significant effect in humans. The main furanocoumarins, xanthotoxin and imperatorin, were also identified and quantified to 280 and 2 µg/tablet, respectively.

Bioactivity-guided isolation of antimicrobial coumarins from Heracleum mantegazzianum Sommier & Levier (Apiaceae) fruits by high-performance counter-current chromatography.

An efficient strategy, based on bioassay-guided fractionation, high-performance liquid chromatography (HPLC), and high-performance counter-current chromatography (HPCCC), was established to purify and evaluate the bioactive compounds from the dichloromethane extract of the fruits of Heracleum mantegazzianum Sommier & Levier (Apiaceae). The quaternary solvent system n-heptane-ethyl acetate-methanol-water (6:5:6:5 v/v) was used in the reversed phase mode. Using this method, in a single run, seven fractions were isolated, among them three were the pure furanocoumarins: pimpinellin, imperatorin, and phellopterin. In order to purify xanthotoxin a more polar system (1:1:1:1 v/v) was further applied. The antimicrobial activity of extract, chromatographic fractions, and single compounds were in the range of MIC = 0.03-1 mg mL(-1). Xanthotoxin may have priority as a compound of further interest based on its antimicrobial activity. For the first time, an extensive antimicrobial study was performed for pimpinellin and phellopterin.

Absence of furanocoumarins in Advantra Z® (Citrus aurantium, bitter orange) extracts.

Grapefruit (Citrus paradisi) juice is known for its ability to alter drug metabolism through inhibition of the cytochrome P450-3A4 (CYP3A4) system, and result in drug-food interactions that may be life threatening. The primary active ingredients in grapefruit responsible for these effects are the furanocoumarins bergapten, bergamottin, and 6',7'-dihydroxybergamottin (DHB). Bergamottin and DHB appear to be the most important in terms of adverse drug interactions. Furanocoumarins are present in the juices and fruits of other Citrus species including C. aurantium (bitter oranges). Bergapten is the predominant furanocoumarin in bitter orange. Bitter orange extracts are widely used in products associated with weight loss, sports performance, and energy production. Questions have been raised about the potential of bitter orange extracts to cause drug interactions. This study examined the furanocoumarin content of four standardized bitter orange extracts (Advantra Z®) by liquid chromatography-mass spectroscopy. The results indicated that the total furanocoumarin content of each of the four extracts was less than 20 µg/g, amounts insufficient to exert significant effects on the metabolism of susceptible drugs in human subjects at the doses commonly used for these extracts.

The effect of pre-treatment and modified atmosphere packaging on contents of phenolic compounds and sensory and microbiological quality of shredded celeriac.

BACKGROUND: The aim of this study was to determine the effect of washing (4 °C, 120 s) or soaking (4 °C, 600 s) of shredded celeriac in tap water on changes in contents of phenolic compounds, including furanocoumarins, and sensory and microbiological quality during 12 days of storage. The product was packaged in air or modified atmosphere containing 2/10/88 kPa O2/CO2/N2. RESULTS: The applied pre-treatment consisting of washing or soaking of shredded celeriac in water resulted in decreases in 8-methoxypsoralen content by approximately 50 and 70% respectively and phenolic content by 30% compared with samples that were not subjected to pre-treatment. During storage of shredded celeriac, a further significant (P ≤ 0.05) reduction in phenolic compounds and an approximately 2.5-fold increase in the total content of furanocoumarins were found. The application of modified atmosphere packaging had a significant effect on the maintenance of good sensory and microbiological quality of the tested product. CONCLUSION: Modified atmosphere packaging of shredded celeriac not subjected to pre-treatment made it possible to obtain a product with good sensory and microbiological quality and the highest content of phenolic compounds. The level of furanocoumarins recorded in the tested product does not constitute a health hazard.

[High-performance liquid chromatography for determination of psoralene, bergapten and apigenin in Ficus hirta Vahl].

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.

In vitro photo-induced cytotoxic activity of Citrus bergamia and C. medica L. cv. Diamante peel essential oils and identified active coumarins.

CONTEXT: The search for innovative therapeutic approaches is gaining more interest in clinical oncology. OBJECTIVE: In the present investigation we reported the chemical profile and the photo-induced cytotoxic activity of two endemic Calabrian Citrus species (Rutaceae): Citrus bergamia Risso & Poit. and Citrus medica L. cv. Diamante. MATERIALS AND METHODS: Essential oils were obtained by hydrodistillation and analyzed by GC and GC/MS. In order to evaluate the cytotoxic activity two melanoma models, such as amelanotic melanoma C32 and malignant melanoma A375, were used. RESULTS: The essential oil of C. bergamia was characterized by limonene, linalyl acetate, gamma-terpinene, linalool and beta-pinene as major components. The most abundant compounds of C. medica cv. Diamante oil were limonene, gamma-terpinene, citral, geranial, beta-pinene and alpha-pinene. Two coumarins, bergapten and citropten, were also identified in C. bergamia and C. medica cv. Diamante, respectively and tested for biological activity. Both C. bergamia and C. medica cv. Diamante oils exhibited a selective interesting activity against the A375 cell line with IC(50) values of 79.3 and 89.1 microg/mL, respectively, after 100 min exposure to UV irradiation. The strong antiproliferative activity demonstrated with bergapten (IC(50) value of 71.3 microg/mL after 20 min of irradiation) was not found with citropten. DISCUSSION AND CONCLUSION: Our study suggested that UV irradiation is effective in activating essential oils and in particular bergapten. This phototoxicity may be considered as a treatment option in some cases of lentigo maligna or lentigo maligna melanoma.

[Simultaneous determination of 5 active components in Fructus Cnidii by HPLC].

OBJECTIVE: To develop an RP-HPLC method for simultaneous determination of 5 constituents in Fructus Cnidii. METHOD: Analysis was performed on an Alltech C18 (4.6 mm x 250 mm, 5 microm) column. The mobile phases were acetonitrile water and acetic acid with gradient elution. The flow rate was 1 mL x min(-1). The monitoring wavelength was 325 nm and 245 nm. The column temperature was 40 degrees C. RESULT: The linear response ranges were 1-20 microg x mL(-1) (r = 0.999 9) for xanthotoxin, 1-20 microg x mL(-1) (r = 0.999 9) for isopimpinellin, 11-20 microg x mL(-1) (r = 0.999 8) for bergapten, 100-1 200 microg x mL(-1) (r = 0.999 7) for imperatorin, 100-2 000 microg x mL(-1) (r = 0.999 9) for osthole. The average recoveries were all above 95%. CONCLUSION: The method is simple, sensitive and accurate with good reproducibility.

Can dietary furanocoumarin ingestion enhance the erythemal response during high-dose UVA1 therapy?

As phototoxic skin reactions caused by psoralen are induced by wavelengths within the UVA1 spectrum, we assessed the potential of the small amount of psoralen in a normal diet to provoke phototoxicity in volunteers with skin types I and II. Threshold erythema was unaffected by ingestion of a 200-g portion of parsnip.

A method for quantitative determination of furanocoumarins in capsules and tablets of phytochemical preparations.

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.

A method for fast determination of psoralens in oral solutions of phytomedicines using liquid chromatography.

A method for sample preparation and analysis by high performance liquid chromatography with UV detection (HPLC-UV) has been developed for routine analysis of psoralen and bergapten, photosensitizing compounds, in oral solutions of phytomedicines employed in Brazil for some illnesses. The linearity, accuracy, the inter- and intra-day precision of the procedure were evaluated. Calibration curves for psoralen and bergapten were linear in the range of 1.0-600.0 microg ml(-1) and 1.0-400.0 microg ml(-1) respectively. The recoveries of the psoralens in the oral solutions analysed were 94.43-99.97%. The percentage coefficient of variation (CV) of the quantitative analysis of the psoralens in the products analysis was within 5%. In inter-equipment study was employed gas chromatography-flame ionization (CG-FID) detection.

2D NMR spectroscopic analyses of archangelicin from the seeds of Angelica archangelica.

A total of six coumarins, bergapten (1), xanthotoxin (2), imperatorin (3), isoimperatorin (4), phellopterin (5) and archangelicin (6), have been isolated from an n-hexane extract of the seeds of Angelica archangelica. The results of comprehensive 2D NMR analyses of archangelicin are discussed.

The furanocoumarins in the roots of Heracleum sibiricum L.

From roots of Heracleum sibiricum L (cow parsnip) collected natural state, a raw crystalline furanocoumarin precipitate was obtained containing four compounds: bergapten, isopimpinellin, pimpinellin, sphondin. The isolated compounds were identified with H1NMR.

Simultaneous determination of furanocoumarins in infusions and decoctions from "Carapiá" (dorstenia species) by high-performance liquid chromatography.

The use of furanocoumarins, which are photosensitizing compounds, combined with exposure to UV-A radiation is a common treatment for vitiligo, psoriasis, and a number of other skin diseases. Although furanocoumarins plus UV-A treatment is highly effective, several studies have shown that exposure to high doses increases the risk to development of cutaneus carcinoma. Several Dorstenia species are used in folk medicine, mainly against skin diseases, because of the presence of biologically active compounds. We present here analysis of the chemical composition of furanocoumarins from infusion and decoction of "Carapiá" (Dorstenia species), which is used in Brazil against several diseases. We have employed high-performance liquid chromatography (HPLC) procedures for the quantitative determination of psoralen, bergapten, and isopimpinellin. The contents of furanocoumarins revealed an insignificant difference between infusion and decoction. Dorstenia tubicina and D. asaroides contained psoralen and bergapten only in the rhizomes, whereas D. vitifolia shows solely isopimpinellin in both rhizomes and aerial parts.

Simple and rapid determination of psoralens in topic solutions using liquid chromatography.

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%.

Bath PUVA--an investigation of the distribution of trioxsalen (TMP) and 8-methoxypsoralen (8-MOP) in bathwater.

Trioxsalen (TMP) bath PUVA avoids the side effects of nausea and headache associated with oral 8-methoxypsoralen (8-MOP) treatment and allows shorter irradiation times that can be advantageous in some patients. However we noted that a number of patients developed unusual patterns of phototoxic burning. We thought that this was related to an uneven distribution of the TMP in the bathwater and for this reason, a study of bath water TMP concentrations achieved using different TMP preparations was undertaken. The distribution of 8-MOP in an 8-MOP bath was also measured for comparison. Our results confirm that an uneven distribution of TMP is achieved using TMP capsules or suspension and would explain our observed patterns of burning. With an ethanolic solution of TMP, or the commercial equivalent Tripsor, or with Puvasoralen-8 (an 8-MOP preparation), a homogeneous psoralen distribution is achieved, and they are therefore preferable for use in bath PUVA.

Rapid identification of some coumarin derivatives in deterpenated citrus peel oil by gas chromatography.

Generally on the gas chromatogram of a volatile essential oil, terpenes, oxygenated compounds and sesquiterpenes appear. With temperature programming, it was shown that some non-volatiles are present with the volatiles. They are simple coumarin (2H-1-benzopyran-2-one) derivatives such as citropten (5,7-dimethoxycoumarin) and furocoumarins (psoralen, 7H-furo[3,2-g][1]benzopyran-7-one) such as bergapten (5-methoxypsoralen), some of which are phototoxic. Terpeneless oils are used in perfumes and cosmetics, so it is important to be able to establish rapidly if they contain phototoxic compounds.

Skin pigmentation as a predictor of minimal phototoxic dose after oral methoxsalen.

BACKGROUND AND DESIGN: Twenty-eight subjects were phototested to determine their erythemal responses to oral methoxsalen with UV-A and UV-B irradiation. Skin pigmentation was measured by skin reflectance at 550 and 660 nm before irradiation. The smallest UV radiation dose to produce erythema (minimal phototoxic dose and minimal erythema dose, respectively) was determined. The serum concentration of methoxsalen was measured at the time of UV-A irradiation. RESULTS: There was a positive correlation between skin pigmentation and both 72-hour minimal phototoxic dose and 24-hour minimal erythema dose. No correlation was demonstrated between methoxsalen serum concentration and minimal phototoxic dose. The combination of skin pigmentation and methoxsalen level did not give a better prediction of minimal phototoxic dose than skin pigmentation alone. CONCLUSIONS: Skin pigmentation measurements can be used to predict the minimal phototoxic and erythema doses. Skin pigmentation measurements are easy to perform and should be included in both phototherapy and photochemotherapy to improve the efficiency and reliability of the treatment.

Characterization and evaluation by 32P-postlabelling of psoralen-type DNA adducts in HeLa cells.

Samples of DNA irradiated at 405 and/or 365 nm in the presence of 8-methoxypsoralen (8-MOP) were analysed via a modified postlabelling assay using three hydrolysis enzymes other than those employed previously. These enzymes (deoxyribonucleaseI, venom phosphodiesterase and alkaline phosphatase) liberated 3'-adducted dinucleotide monophosphate instead of the 5'-modified dinucleotide monophosphate normally obtained. The first separation chromatography (D1) of samples irradiated in the presence of 8-MOP showed a single spot above the origin, and the next separation (D2) resolved this spot into two components (spots I and II). Double irradiation experiments in which samples of DNA were first irradiated at 405 nm before being irradiated at 365 nm showed that spot II could be transformed into spot I. The use of 6,4,4'-trimethylangelicin, which induced only photomonoadducts under UVA irradiation, gave only spot II. These two results indicated that spots I and II were respectively due to interstrand cross-links and monoadducts. Dose-effect experiments showed that spots I and II were dose dependent, and low-dose irradiations permitted us to measure one interstrand cross-link and two monoadducts per 10(8) base pairs.