Simultaneous determination of multiple components in rat plasma and pharmacokinetic studies at a pharmacodynamic dose of Xian-Ling-Gu-Bao capsule by UPLC-MS/MS.

Xian-Ling-Gu-Bao capsule (XLGB) is an effective traditional Chinese medicine prescription (TCMP) that is used for the prevention and treatment of osteoporosis in China. A rapid, simple, efficient and stable method based on UPLC-MS/MS technology was developed for simultaneous determination of multiple components of XLGB in rat plasma. Mass spectrometric detection was performed in multiple reaction monitoring (MRM) mode with electrospray ionization (ESI). For twenty-one selected quantitative prototypes, all calibration curves showed favourable linearity (r>0.9932) in linear ranges. The lower limits of quantification (LLOQs) were 2 ng/mL for psoralen (PL), 2.5 ng/mL for asperosaponin VI (AS), 1 ng/mL for isopsoralen (IPS) and sweroside (SW), 0.5 ng/mL for magnoflorine (MA), bavachinin (BVN), tanshinone IIA (TA), timosaponin BII (TBII) and icaritin (ICT), 0.1 ng/mL for epimedin B (EB) and epimedin C (EC), 0.05 ng/mL for icariin (IC), isobavachalcone (IBC), psoralidin (PD), bavachin (BV), bavachalcone (BC), epimedin A (EA) and isobavachin (IBV), 0.02 ng/mL for neobavaisoflavone (NEO) and icariside I (ICI) and 0.01 ng/mL for icariside II (ICII). The intra-day and inter-day (low, medium, high) precision (relative standard deviation) for all analytes was less than 8.63%, and the accuracies (as relative error) were in the range of -12.45% to 8.91%. Extraction recoveries and matrix effects of analytes and IS were acceptable. All analytes were stable during the assay and storage in plasma samples. The validated method was successfully applied to the pharmacokinetics (PK) studies of the twenty-one prototypes at pharmacodynamic doses (0.3 and 1 g/kg/day). In addition, dynamic profiles of 28 metabolites (phase II conjugates: 23 glucuronide conjugates, 2 sulfate conjugates and 3 glucuronide or sulfate conjugates) were also monitored by their area/IS area-time curves. As a result, coumarins, prenylated flavonoids from Psoraleae Fructus, alkaloids and prenylated flavonol glycosides from Epimedii Herba, and iridoid glycosides, triterpenoid saponins from Dipsaci Asperoidis Radix were considered to be the key effective substances of XLGB due to their high exposure and appropriate pharmacokinetic features. This is the first report to reveal pharmacodynamic ingredients by a reversed pharmacodynamic (PD) - pharmacokinetics (PK) study.

Evaluation of the Absorption Behavior of Main Component Compounds of Salt-Fried Herb Ingredients in Qing'e Pills by Using Caco-2 Cell Model.

Qing'e Pills is a Chinese traditional herbal product, which is often used to strengthen muscles and bones in TCM (traditional Chinese Medicine) practice. Its two main component herbs, namely, Cortex Eucommiae and Fructus Psoraleae are both required to be salt-fried according to TCM theory. We have evaluated the effects of salt-frying treated herbs on Caco-2 cell uptake behavior for those active ingredients of Qing'e Pills. By investigating of various variables, including MTT, temperature, inhibitors, pH, salt concentration and herb processing methods, we tried to clarify whether the salt-processing on herbs was necessary or not. Results showed that, compared to other processing methods, the salt-frying process significantly (p < 0.01) enhanced the absorption of effective components of Qing'e Pills. The way that psoralen, isopsoralen, psoralenoside and geniposide acid entered Caco-2 cells at low concentrations was via passive diffusion. These components were not substrates of P-glycoprotein. It demonstrated that the salt-frying process not only enhanced the concentration of active components in herb extract, but also changed their absorption behaviors. Nevertheless, the mechanism of absorption behavior changing needs to be further investigated.

[Determination and pharmacokinetics of main components for Psoralea corylifolia-Myristica fragrants drug pair by using UPLC-MS/MS].

To conduct multiple-reaction monitoring(MRM) quantitative analysis with ultra-high performance liquid chromatography coupled with mass spectrometry method(UPLC-MS/MS), determine the concentrations of psoralen, isopsoralen, bakuchiol and dehydrodiisoeugenol in plasma under positive iron mode with chloramghenicol as internal standard, and investigate the pharmacokinetics process of the main components before and after oral administration of drug pair Psoralea corylifolia -Myristica fragrants. Thirty-six SD rats were randomly divided into three group(A, B, C) and received P. corylifolia extract, P. corylifolia-M. fragrants extract, and M. fragrants extract respectively by intragastric administration. The plasma samples were collected at different time points. In the plasma samples, psoralen, isopsoralen, bakuchiol and dehydrodiisoeugenol showed good linear relationship within concentration rages of 0.098 125 to 39.25, 0.084 37 to 33.75, 0.046 875 to 18.75, and 0.11 to 2.2 mg•L⁻¹ respectively. The precision and stability results showed that the determination method of plasma concentration for such compositions was stable and reliable. The pharmacokinetic parameters obtained by DAS 2.0 showed varying differences before and after compatibility. According to the experimental results, the compatibility of P. corylifolia and M. fragrants can significantly impact the pharmacokinetic process of main components, expand their distribution and accelerate their metabolism and elimination in vivo.

Evaluation of the influence of salt processing on pharmacokinetics of psoralen and isopsoralen in Psoralea corylifolia L.

A sensitive, specific and rapid ultra-high-pressure liquid chromatography tandem mass spectrometry (UHPLC-MS/MS) method has been developed to investigate pharmacokinetic properties of psoralen and isopsoralen, two compounds isolated from raw/salt-processed fruit of Psoralea corylifolia L. UHPLC-MS/MS was used with positive ion electrospray. The mobile phase was composed of acetonitrile and 0.1% formic acid aqueous solution and a gradient elution program at flow rate of 0.3 mL/min was applied. Multiple reaction monitoring mode was used for the quantification of psoralen, isopsoralen ([M + H](+) m/z 187.0 → m/z 131.0) and scoparone (m/z 207.0 → m/z 151.1). Scoparone served as an internal standard. The method was fully validated for its sensitivity, selectivity, stability, matrix effect and extraction recovery. The obtained results showed that salt-processed Buguzhi significantly promoted the absorption of psoralen and isopsoralen, and increased the bioavailability of these compounds.

A UPLC-MS/MS method for in vivo and in vitro pharmacokinetic studies of psoralenoside, isopsoralenoside, psoralen and isopsoralen from Psoralea corylifolia extract.

ETHNOPHARMACOLOGICAL RELEVANCE: The dried fruit of Psoralea corylifolia L. has been used to prevent and treat vitiligo, osteoporosis, arthralgia and asthma in Traditional Chinese Medicine for some 1600 years. Psoralen (P), isopsoralen (IP), psoralenoside (PO) and isopsoralenoside (IPO) are the major coumarins and coumarin-related benzofuran glycosides in Psoraleae Fructus, which have been reported to show estrogen-like activity, osteoblastic proliferation accelerating activity, antitumor effects and antibacterial activity. The first aim of this study is to develop a rapid, sensitive and selective ultra performance liquid chromatography tandem mass spectrometry (UPLC-MS/MS) approach for simultaneous determination of PO, IPO, P and IP in rat plasma and samples collected from in vitro incubation experiments. The second aim is to investigate the pharmacokinetic properties of PO, IPO, P and IP after oral administration of Psoralea corylifolia extract (PCE) to rats. The third aim is to confirm the biotransformation of PO to P or IPO to IP under gastrointestinal conditions. MATERIALS AND METHODS: A UPLC-MS/MS method with a C18 column and a mobile phase of methanol-0.1% aqueous formic acid was validated according to the criteria in FDA guidelines about bioanalytical method, which was developed to investigate the pharmacokinetic behavior of PO, IPO, P and IP from PCE and the metabolic pathways of PO to P or IPO to IP. RESULTS: The criteria for establishment of a new UPLC-MS/MS method including selectivity, linearity, accuracy, precision, extraction recovery, matrix effect and stability were validated. This method was successfully applied to the quantitative determination of PO, IPO, P and IP in biological samples collected from both in vitro incubations and in vivo rat experiments. After oral administration of PCE to rat, pharmacokinetic parameters of these four compounds indicated that in vivo biotransformation may occur between PO and P or IPO and IP. Purified benzofuran glycosides fraction (PBGF), containing only PO and IPO, was orally administered to rats to further confirm the biotransformation of PO to P or IPO to IP under gastrointestinal conditions. An in vitro incubation study elucidated that PO and IPO were metabolized to P and IP by intestinal microflora through de-glucosylation. CONCLUSIONS: This paper developed a rapid, sensitive and selective UPLC-MS/MS method for simultaneous determination of PO, IPO, P and IP from PCE in biological samples, and investigated on their comprehensive in vivo and in vitro pharmacokinetic studies. These obtained results showed that the metabolism by intestinal bacteria plays an important role in pharmacological effects of orally administered PCE.

Simultaneous determination of scopoletin, psoralen, bergapten, xanthotoxin, columbianetin acetate, imperatorin, osthole and isoimperatorin in rat plasma by LC-MS/MS for pharmacokinetic studies following oral administration of Radix Angelicae Pubescentis extract.

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.

[Comparative study of absorption kinetics in intestines of rats on Xianlinggubao capsule prepared by different technologies].

OBJECTIVE: To study the characteristics of intestinal absorption of psoralen and isopsoralen of Xianlinggubao capsule, and compare the absorption of Xianlinggubao capsule prepared by different technologies. METHOD: Non everted gut sac method was applied to investigate the influence of absorption sites and drug concentration on psoralen and isopsoralen absorption, which were determined by HPLC. RESULT: Although the absorption rate constants of psoralen and isopsoralen in duodenum were more than that in jejunum and ileum, there was no significance difference between them. The absorption rate constants of psoralen kept at the same level when the concentrations of drug solution were at middle and low level, while the absorption rate constant at high level was absolutely lower than them (P < 0.05). The results of isopsoralen were the same as psoralen's. CONCLUSION: Intestinal absorption of psoralen and isopsoralen may be affected by the dissolution. The absorption rate constants of psoralen and isopsoralen in new Xianlinggubao capsules are higher. The absorptions of active components absorption has significant difference in different preparation processes of Xianlinggubao capsule.

Pharmacokinetics, tissue distribution and excretion of coumarin components from Psoralea corylifolia L. in rats.

Coumarin components from Psoralea corylifolia L. are novel drugs in which psoralen and isopsoralen are the active components. The pharmacokinetics, tissue distribution and excretion of the two compounds were studied by liquid chromatography-tandem mass spectrometry after intravenous administration to Wistar rats. The elimination half-lives of psoralen and isopsoralen were 4.88 and 5.35 h. After dosing, the area under the curves of the tissues decreased in the following order: liver > lung > heart > kidney > spleen > brain for psoralen; and kidney > lung > liver > heart > spleen > brain for isopsoralen. After dosing, 51.27% of psoralen and 56.25% of isopsoralen were excreted as prototype, and urine was the major excretion route. In addition, the pharmacokinetics of psoralen and isopsoralen after oral administration to Wistar rats were also studied. The elimination half-lives of psoralen and isopsoralen were 4.13 and 5.56 h, and their relative bioavailabilities were 61.45% and 70.35%. Overall, the results show that coumarin components from P. corylifolia L. have high oral bioavailability, they are rapidly and widely distributed into tissues after intravenous administration, but they are slowly cleared and excreted.

Simultaneous quantification of psoralen and isopsoralen in rat plasma by ultra-performance liquid chromatography/tandem mass spectrometry and its application to a pharmacokinetic study after oral administration of Haigou Pill.

A rapid, specific and sensitive ultra-performance liquid chromatography/tandem mass spectrometry (UPLC/MS/MS) method has been established for simultaneous quantitation of psoralen and isopsoralen in rat plasma. Plasma samples were pretreated by direct protein precipitation with acetonitrile. Chromatographic separations were performed on an ACQUITY UPLC BEH C(18) column (50mmx2.1mm, i.d., 1.7microm) at 35 degrees C with a linear gradient of acetonitrile and 0.1% formic acid in water at a flow rate of 0.3mL/min. The two isomers were satisfactorily separated (R=1.7) with a runtime of 4min. Psoralen, isopsoralen, and the internal standard (IS) furazolidone were ionized with an APCI source operated in positive ion mode. The MS/MS transitions used for monitoring were at m/z 187.0-->130.9 for psoralen and isopsoralen, and m/z 225.9-->121.9 for IS. Calibration curve was linear over the concentration range of 1-500ng/mL with the lower limit of quantitation of 1ng/mL for both isomers. The mean extraction recoveries were 78.5+/-6.7% and 81.9+/-8.0% for psoralen and isopsoralen, respectively. The intra- and inter-day precisions were less than 5.6% and 5.2%, and the accuracy was within +/-2.1% for both isomers. No matrix effect was observed in this method. Psoralen and isopsoralen were stable during all storage, pretreatment and analytical periods. The validated method has been successfully applied to a pharmacokinetic study of psoralen and isopsoralen after oral administration of Haigou Pill to rats.

In vivo pharmacokinetics comparisons of icariin, emodin and psoralen from gan-kang granules and extracts of herba Epimedii, Nepal dock root, Ficus hirta yahl.

ETHNOPHARMACOLOGICAL RELEVANCE: Gan-kang granules (GKG) contains the extracts from eight Chinese herbs and is a traditional Chinese composite prescription for treatment of hepatitis B. Icariin, emodin and psoralen are main effective ingredients of the medicine. AIM OF THE STUDY: In this research, pharmacokinetic comparisons of icariin, emodin and psoralen from the extracts of herba Epimedii, Nepal dock root and Ficus hirta yahl, and GKG were conducted. MATERIALS AND METHODS: At different time points (0.25, 0.5, 0.75, 1, 1.5, 2, 4, 6, 8, 12 and 24 h) after administration, the concentrations of icariin, emodin and psoralen in rat plasma were determined by HPLC-UV, and main pharmacokinetic parameters were estimated. RESULTS: The pharmacokinetic parameters of icariin, emodin and psoralen in GKG were elevated comparing with those of herb extracts. CONCLUSIONS: Three HPLC-UV methods were developed successfully for the analysis of icariin, emodin and psoralen in SD rat plasma. Some ingredients in GKG may increase the dissolution and absorption, and improve bioavailability of icariin, emodin and psoralen in rats.

Preclinical safety of a nucleic acid-targeted Helinx compound: a clinical perspective.

Helinx technology (Cerus Corp, Concord, CA) uses amotosalen HCl (S-59) and ultraviolet A (UVA) light in an ex vivo photochemical treatment (PCT) to inactivate viruses, bacteria, and leukocytes in platelet concentrates while preserving therapeutic function. A comprehensive preclinical safety program was conducted, which included carcinogenicity, single-dose and multiple-dose (up to 13 weeks' duration) toxicity, safety pharmacology (central nervous system [CNS], renal, and cardiovascular), reproductive toxicity, genotoxicity, vein irritation, phototoxicity, and toxicokinetic testing. The results of the toxicokinetic analyses indicated that the test articles provided large multiples of the clinical exposure to S-59, whether the comparison was based on dose, maximum plasma concentration, or area under the concentration-time curve. No specific target organ toxicity, reproductive toxicity, or carcinogenicity was observed. S-59 and/or PCT formulations demonstrated CNS toxicity, electrocardiographic (ECG) effects, and phototoxicity at supraclinical doses. On the basis of the extremely large safety margins, the CNS and ECG observations (at >30,000-fold the expected clinical exposure) as well as the results of genotoxicity and phototoxicity studies are not considered to be of toxicological relevance. The results of an extensive series of studies have thus demonstrated no toxicologically relevant effects of platelets treated with Helinx technology.

Inhibition of rat brain monoamine oxidase activities by psoralen and isopsoralen: implications for the treatment of affective disorders.

Psoralen and isopsoralen, furocoumarins isolated from the plant Psoralea corylifolia L., were demonstrated to exhibit in vitro inhibitory actions on monoamine oxidase (MAO) activities in rat brain mitochondria, preferentially inhibiting MAO-A activity over MAO-B activity. This inhibition of enzyme activities was found to be dose-dependent and reversible. For MAO-A, the IC50 values are 15.2 +/- 1.3 microM psoralen and 9.0 +/- 0.6 microM isopsoralen. For MAO-B, the IC50 values are 61.8 +/- 4.3 microM psoralen and 12.8 +/- 0.5 microM isopsoralen. Lineweaver-Burk transformation of the inhibition data indicates that inhibition by both psoralen and isopsoralen is non-competitive for MAO-A. The Ki values were calculated to be 14.0 microM for psoralen and 6.5 microM for isopsoralen. On the other hand, inhibition by both psoralen and isopsoralen is competitive for MAO-B. The Ki values were calculated to be 58.1 microM for psoralen and 10.8 microM for isopsoralen. These inhibitory actions of psoralen and isopsoralen on rat brain mitochondrial MAO activities are discussed in relation to their toxicities and their potential applications to treat affective disorders.

Pharmacokinetic and toxicology assessment of INTERCEPT (S-59 and UVA treated) platelets.

The pathogen inactivation process developed by Cerus and Baxter Healthcare Corporations uses the psoralen, S-59 (amotosalen) in an ex vivo photochemical treatment (PCT) process to inactivate viruses, bacteria, protozoans, and leukocytes in platelet concentrates and plasma. Studies were performed by intravenous infusion of S-59 PCT formulations +/- compound adsorption device (CAD) treatment and with non-UVA illuminated S-59, using doses that were multiples of potential clinical exposures. The studies comprised full pharmacokinetic, single- and repeated-dose (up to 13 weeks duration) toxicity, safety pharmacology (CNS, renal, and cardiovascular), reproductive toxicity, genotoxicity, carcinogenicity testing in the p53(+/-) mouse, vein irritation, and phototoxicity. No specific target organ toxicity (clinical or histopathological), reproductive toxicity, or carcinogenicity was observed. S-59 and/or PCT formulations demonstrated CNS, ECG, and phototoxicity only at supraclinical doses. Based on the extremely large safety margins (>30,000-fold expected clinical exposures), the CNS and ECG observations are not considered to have any toxicological relevance. Additionally, after a complete assessment, mutagenicity and phototoxicity results are not considered relevant for the proposed use of INTERCEPT platelets. Thus, the results of an extensive series of in vitro and in vivo studies have not demonstrated any toxicologically relevant effects of platelet concentrates prepared by the INTERCEPT system.

[A study on nasal absorption of psoralen and isopsoralen in Psoralea corylifolia L].

OBJECTIVE: To investigate the nasal absorption regularities of psoralen and isopsoralen of different concentrations. METHOD: Building an experimental model of rat in situ nasal recirculation and determining the contents of psoralen and isopsoralen by HPLC. RESULT: The nasal absorption of psoralen and isopsoralen fitted in with zero order kinetics, getting saturated with the increase of concentration. CONCLUSION: A suitable concentration is necessary for the preparation of nasal remedies psoralen and isopsoralen.