Sanguinarine metabolism and pharmacokinetics study in vitro and in vivo.

Sanguinarine (SA) is a benzo[c] phenanthridine alkaloid which has a variety of pharmacological properties. However, very little was known about the pharmacokinetics of SA and its metabolite dihydrosanguinarine (DHSA) in pigs. The purpose of this work was to study the intestinal metabolism of SA in vitro and in vivo. Reductive metabolite DHSA was detected during incubation of SA with intestinal mucosa microsomes, cytosol, and gut flora. After oral (p.o.) administration of SA, the result showed SA might be reduced to DHSA in pig intestine. After i.m. administration, SA and DHSA rapidly increased to reach their peak concentrations (Cmax , 30.16 ± 5.85, 5.61 ± 0.73 ng/ml, respectively) at 0.25 hr. Both compounds were completely eliminated from the plasma after 24 hr. After single oral administration, SA and DHSA rapidly increased to reach their Cmax (3.41 ± 0.36, 2.41 ± 0.24 ng/ml, respectively) at 2.75 ± 0.27 hr. The half-life (T1/2 ) values were 2.33 ± 0.11 hr and 2.20 ± 0.12 hr for SA and DHSA, respectively. After multiple oral administration, the average steady-state concentrations (Css ) of SA and DHSA were 3.03 ± 0.39 and 1.42 ± 0.20 ng/ml. The accumulation indexes for SA and DHSA were 1.21 and 1.11. The work reported here provides important information on the metabolism sites and pharmacokinetic character of SA. It explains the reasons for low toxicity of SA, which is useful for the evaluation of its performance.

Synthesis and biological evaluation of 5-aminoethyl benzophenanthridone derivatives as DNA topoisomerase IB inhibitors.

DNA topoisomerase IB (TOP1) regulates DNA topological structure in many cellular metabolic processes and is a validated target for development of antitumor agents. Our previous study revealed that the benzophenanthridone scaffold is a novel chemotype for the discovery of TOP1 inhibitors. In this work, a series of novel 5-aminoethyl substituted benzophenanthridone derivatives have been synthesized and evaluated for TOP1 inhibition and cytotoxicity. Compound 12 exhibits the most potent TOP1 inhibition (+++) and cytotoxicity in human cancer cell lines with GI50 values at nanomolar concentration range. 12 induces the cellular TOP1cc formation and DNA damage, resulting in HCT116 cell apoptosis. The pharmacokinetics, acute toxicity and antitumor efficiency in vivo of 12 were also studied.

Pharmacokinetics of sanguinarine, chelerythrine, and their metabolites in broiler chickens following oral and intravenous administration.

Sanguinarine (SA) and chelerythrine (CHE) are the main active components of the phytogenic livestock feed additive, Sangrovit®. However, little information is available on the pharmacokinetics of Sangrovit® in poultry. The goal of this work was to study the pharmacokinetics of SA, CHE, and their metabolites, dihydrosanguinarine (DHSA) and dihydrochelerythrine (DHCHE), in 10 healthy female broiler chickens following oral (p.o.) administration of Sangrovit® and intravenous (i.v.) administration of a mixture of SA and CHE. The plasma samples were processed using two different simple protein precipitation methods because the parent drugs and metabolites are stable under different pH conditions. The absorption and metabolism of SA following p.o. administration were fast, with half-life (t1/2 ) values of 1.05 ± 0.18 hr and 0.83 ± 0.10 hr for SA and DHSA, respectively. The maximum concentration (Cmax ) of DHSA (2.49 ± 1.4 µg/L) was higher that of SA (1.89 ± 0.8 µg/L). The area under the concentration vs. time curve (AUC) values for SA and DHSA were 9.92 ± 5.4 and 6.08 ± 3.49 ng/ml hr, respectively. Following i.v. administration, the clearance (CL) of SA was 6.79 ± 0.63 (L·h-1 ·kg-1 ) with a t1/2 of 0.34 ± 0.13 hr. The AUC values for DHSA and DHCHE were 7.48 ± 1.05 and 0.52 ± 0.09 (ng/ml hr), respectively. These data suggested that Sangrovit® had low absorption and bioavailability in broiler chickens. The work reported here provides useful information on the pharmacokinetic behavior of Sangrovit® after p.o. and i.v. administration in broiler chickens, which is important for the evaluation of its use in poultry.

Biotransformation and tissue distribution of protopine and allocryptopine and effects of Plume Poppy Total Alkaloid on liver drug-metabolizing enzymes.

In this study, the biotransformation in the plasma, urine and feces of rats following oral administration of protopine (PRO) and allocryptopine (ALL)were explored using HPLC-QqTOF MS. An HPLC-MS/MS method for the determination of tissues was developed and applied to the tissue distribution study in rats following intragastric administration of Plume Poppy Total Alkaloid for 3 weeks. A total of ten PRO metabolites and ten ALL metabolites were characterized in rats in vivo. Among these metabolites, six PRO metabolites and five ALL metabolites were reported for the first time. The predicated metabolic pathways including ring cleavage, demethylation following ring cleavage, and glucuronidation were proposed. The low-concentration residue of PRO and ALL in various tissues was detected at 24 h and 48 h after dosing, which indicated that both compounds could be widely distributed in tissues and exist as low levels of residue. The activities of erythromycin N-demethylase, aminopyrine N-demethylase and NAD (P)H quinone oxidoreductase in female rats can be induced post-dose, but these activities were inhibited in male rats. The proposed biotransformation and residues of PRO and ALL and their effects on enzymes may provide a basis for clarifying the metabolism and interpreting pharmacokinetics.

Enrichment of chelidonine from Chelidonium majus L. using macroporous resin and its antifungal activity.

Chelidonium majus L. (greater celandine) has been used as an herbal medicine for several centuries. This study investigated an efficient method to purify chelidonine from the extract of C. majus L. using macroporous adsorption resins and evaluated the antifungal activity of chelidonine against Botryosphaeria dothidea as a model strain. Static adsorption and desorption tests revealed that D101 was the optimal resin for chelidonine purification. Pseudo-second-order kinetics model and Freundlich equation model were the most suitable for evaluating the endothermic and spontaneous adsorption processes of chelidonine on D101. Dynamic adsorption and desorption tests on D101 columns showed that the concentration of chelidonine increased 14.16-fold, from 2.67% to 37.81%, with the recovery yield of 80.77%. The antifungal activity of enriched chelidonine products was studied with B. dothidea. The results showed that the EC50 of crude extracts, enriched chelidonine products, and chelidonine standard against B. dothidea were 3.24mg/mL, 0.43mg/mL, and 0.77mg/mL, respectively. The result of antifungal activity test showed that chelidonine had the potential to be a useful antifungal agent. Moreover, the enrichment method of chelidonine was highly efficient, low cost, and harmless to the environment for industrial applications.

Chelerythrine and Fe3O4 Loaded Multi-Walled Carbon Nanotubes for Targeted Cancer Therapy.

The work focused on manufacturing improved drug loaded multifunctional magnetic nanoparticles that can overcome the relative non-specificity and potential side-effects of some chemotherapeutic drugs to healthy tissues. A new drug delivery system, Chelerythrine (CHE) and Fe3O4 loaded multi-walled carbon nanotubes (Fe3O4/MWNTs-CHE nanocomposites) that can target hepatocytes when treating malignant tumors, was prepared through a simple adsorption method. The formulation and structure of the Fe3O4/MWNTs-CHE nanocomposites were characterized by vibrating sample magnetometer (VSM), Fourier Transform infrared spectroscopy (FTIR) and Scanning Electron Microscopy (SEM). The cytotoxicity and anti-proliferation effect from the prepared nanocomposites were in vitro tested on human hepatocarcinoma HepG2 and normal liver LO2 cell lines. The results showed the saturated magnetization of Fe3O4/MWNTs-CHE nanocomposites could reach to 45.4O3 emu/g, and the in vitro CHE release behavior exhibited a biphasic release pattern. Moreover, the in vitro cytotoxicity studies revealed that the Fe3O4/MWNTs-CHE nanocomposites showed an efficient inhibition rate to HepG2 cell line and exhibited a lower cytotoxicity to LO2 cell line in comparison to the native CHE. Therefore, the multifunctional Fe3O4/MWNTs-CHE nanocomposites should be a useful and promising candidate for treatment of malignant tumors.

Role of OCT2 and MATE1 in renal disposition and toxicity of nitidine chloride.

BACKGROUND AND PURPOSE: Nitidine chloride (NC), a benzophenanthridine alkaloid, has various biological properties including anticancer and analgesic activities. The aim of the present study was to evaluate the role of organic cation transporter 2 (OCT2) and multidrug and toxin extrusion 1 (MATE1) in the renal disposition and nephrotoxicity of NC. EXPERIMENTAL APPROACH: MDCK cells stably expressing human OCT2 and/or hMATE1 were used to investigate the OCT2- and MATE1-mediated transport of NC. In addition, the accumulation of NC and its potential toxicity were studied in rat primary-cultured proximal tubular (rPCPT) cells and in rats in vivo. KEY RESULTS: NC was found to be a high-affinity substrate of both OCT2 and MATE1 with high cytotoxicity in MDCK-hOCT2/hMATE1 and MDCK-hOCT2 compared to mock cells. The OCT2 inhibitors, cimetidine and (+)-tetrahydropalmatine ((+)-THP), significantly reduced NC accumulation and cytotoxicity in MDCK-hOCT2, MDCK-hOCT2/hMATE1 and rPCPT cells. Severe kidney damage with high levels of blood urea nitrogen and lactate dehydrogenase (LDH), reduced levels of alkaline phosphatase (ALP) and pathological changes were found in rats after 20 days of successive i.v. doses of NC (5 mg·kg(-1) ·day(-1) ). Concomitantly, the concentration of NC in the kidney reached similar high levels at 2 h after the last dose of the 20 day treatment as those observed at 0.5 h after a single i.v. dose of 5 mg·kg(-1) . CONCLUSIONS AND IMPLICATIONS: Our data indicate that NC-induced nephrotoxicity might be mainly attributed to OCT2-mediated extensive renal uptake and weak tubular secretion by MATE1.

Antibacterial efficacy and pharmacokinetic evaluation of sanguinarine in common carp (Cyprinus carpio) following a single intraperitoneal administration.

Sanguinarine (SA), with antimicrobial and antiparasitic activities against fish pathogens, exhibits great potential commercial use in aquaculture. However, little information on pharmacokinetics of SA restricts further application in aquaculture. In this study, pharmacokinetics of SA in common carp (Cyprinus carpio) following a single intraperitoneal administration [10 mg kg(-1) BW (body weight)] was evaluated by high-performance liquid chromatography (HPLC). The peak concentration (Cmax ) of SA in kidney was 11.8 µg g(-1) , which was higher than in other tissues and plasma. The terminal half-life in fish tissue and plasma was as follows: 42.3 h (kidney) > 37.2 h (liver) > 20.1 h (gill) > 18.8 h (muscle) > 10.9 h (spleen) > 10.0 h (plasma). Additionally, we determined the bacterial loads in tissues of common carp infected with Aeromonas hydrophila after i.p. administration of SA at 0, 5, 10 and 20 mg kg(-1) BW. The results showed that i.p. administration of SA at 10 mg kg(-1) BW significantly enhanced antibacterial efficacy against A. hydrophila, where the antibacterial ratio in the gill, kidney, spleen and liver on day 5 was 95.13%, 93.33%, 90.09% and 92.82%, respectively. Overall, these results suggested the potential of SA to treat A. hydrophila infection in common carp farming industry.

Simultaneous quantitative determination of sanguinarine, chelerythrine, dihydrosanguinarine and dihydrochelerythrine in chicken by HPLC-MS/MS method and its applications to drug residue and pharmacokinetic study.

A specific and reliable HPLC-MS/MS method was developed and validated for simultaneously determination of sanguinarine, chelerythrine and their metabolites (dihydrosanguinarine and dihydrochelerythrine) in chicken tissue for the first time. This is important because these compounds are related to the use of a naturally occurring and novel feed additive with many benefits, but the levels of these compounds must be strictly controlled. The compounds were extracted by acetonitrile and 1% HCl-methanol solution successively and then separated on a C18 column. A triple-quadrupole mass spectrometer equipped with an electrospray ionization (ESI) source was used for detection. Quantification was performed using multiple reaction monitoring with positive mode. The method was validated in terms of specificity, linearity, precision, accuracy and stability. The calibration curves were linear over the concentration range of 0.5-100.0ng/g for sanguinarine, 0.5-100.0ng/g for chelerythrine, 0.2-100.0ng/g for dihydrosanguinarine and 0.1-100ng/g for dihydrochelerythrine, respectively. All of the recovery rates of the four analytes were over 85%. The RSD of intra-day and inter-day precision was less than 5.0%, and the relative error was all within 12.0%. This validated method has been successfully applied to assess the drug residue and metabolite residue characteristics of sanguinarine and chelerythrine in chicken tissue after oral administration of the extracts of Macleaya cordata (Willd.) R. Br, and to investigate the pharmacokinetic parameters of sanguinarine and dihydrosanguinarine in chicken plasma.

Pharmacokinetic and anti-inflammatory effects of sanguinarine solid lipid nanoparticles.

The sanguinarine (SG) was studied for its pharmacokinetic and anti-inflammatory activities with prepared solid lipid nanoparticles (SLNs). The sanguinarine solid lipid nanoparticles (SG-SLNs) were prepared by film-ultrasonic dispersion method and the entrapment efficiency of SG was higher at 75.6 %. The drug release profile of SG was examined in pH 7.4 PBS and 85 % of the SG loaded in SLNs was gradually released during 24 h. We used mice endotoxin shock model which was induced by lipopolysaccharide (1 mg/kg) to examine the anti-inflammatory function of SG-SLNs. Healthy Kunming mice were administered orally with saline, SG (10 mg/kg), and SG-SLNs (10 mg/kg), respectively, at 12 and 1 h before lipopolysaccharide (LPS) injection. Mice were sacrificed at 1 and 6 h, respectively, and blood was collect through the venous sinus to access inflammatory mediators. Pharmacokinetic studies proved that the AUC(0→24) and C(max) of SG-SLNs were significantly increased compared that of SG. SG-SLNs revealed significant anti-inflammatory effects through inhibition of LPS-induced tumor necrosis factor-alpha level, interleukin 6 level, and nitric oxide production in serum. Therefore, it can be concluded that SG-SLNs led to a better oral bioavailability.

Synergistic antimicrobial activity of combinations of sanguinarine and EDTA with vancomycin against multidrug resistant bacteria.

A combination of antimicrobial drugs has a potential to overcome multidrug resistant pathogens. In our study we tested the combination of an antimicrobial DNA-intercalating alkaloid (sanguinarine), a chelator (EDTA) with a standard antibiotic (vancomycin), i.e. drugs, which differ in their mode of action. The antibacterial activities of individual substances and of two-drug and three-drug combinations were evaluated for 34 strains of Gram-positive and Gramnegative bacteria (among them 23 clinical isolates) which are not sensitive for vancomycin. MIC and MBC values were determined for each drug individually. Sanguinarine demonstrated a strong activity against all the strains; its activity was comparable to that of antibiotics (MIC = 0.5 - 128 µg/ml). Time kill pharmacokinetics were studied for different concentrations of sanguinarine. A sanguinarine concentration of 16 x MIC was bactericidal against both Gram-positive and Gram-negative strains within 4 to 6 h of incubation. EDTA has only bacteriostatic activity against both Gram-positive and Gram-negative bacteria. As expected, vancomycin is active against Gram-positive bacteria (MIC = 0.125 - 16 µg/ml) but much weaker against Gram-negative bacteria (MIC = 4 - 128 µg/ml). Using the checkerboard design, two- and threedrug combinations were evaluated. Additive and synergistic effects were recorded for all sanguinarine + EDTA and sanguinarine + EDTA + vancomycin combinations against Gram-negative bacteria. Time kill assays indicated that only the combination of 1 x MIC sanguinarine + 1 x MIC EDTA + 1 x MIC vancomycin resulted in a synergistic interaction against MRSA. In the combination assays Gram-negative bacteria became sensitive for vancomycin. More experiments are needed to demonstrate that such a combination strategy also works under in vivo conditions and is clinically relevant.

Mass spectrometric investigation of chelerythrine and dihydrochelerythrine biotransformation patterns in human hepatocytes.

The quaternary benzo[c]phenanthridine alkaloids (QBAs) are an important subgroup of plant secondary metabolites. Their main representatives, sanguinarine (SG) and chelerythrine (CHE), have pleiotropic biological effects and a wide spectrum of medicinal applications. The biotransformation of SG and CHE has only been partially studied while subsequent oxidative transformation of their dihydro derivates, the main metabolites, is practically unknown. The aim of this study was to characterize the biotransformation of CHE and dihydrochelerythrine (DHCHE) in detail, with respect to their more extensive biotransformation than SG. Phase I as well as phase II biotransformation of both compounds was examined in human hepatocyte suspensions. Liquid chromatography with electrospray-quadrupole time-of-flight mass spectrometry (LC-ESI-QqTOF MS) was used for analysis of the metabolites. Using the LC-ESI-QqTOF MS method, we analyzed and then suggested the putative structures of 11 phase I and 5 phase II metabolites of CHE, and 11 phase I and 6 phase II metabolites of DHCHE. For the most abundant metabolites of CHE, DHCHE and O-demethylated DHCHE, their cytotoxicity on primary cultures of human hepatocytes was analyzed. Both metabolites were nontoxic up to 50µM concentration and this indicates decreasing toxic effects for CHE biotransformation products, i.e. DHCHE and O-demethylated DHCHE.

Poly (lactide-co-glycolide) nano-encapsulation of chelidonine, an active bioingredient of greater celandine (Chelidonium majus), enhances its ameliorative potential against cadmium induced oxidative stress and hepatic injury in mice.

This study evaluates the possible protective potentials of chelidonine and its poly lactide-co-glycolide (PLGA) encapsulated nano-form against cadmium chloride (CdCl2) induced oxidative stress and hepatotoxicity in mice, ex vivo and in vivo. Acute exposure to CdCl2 (1.0 mg/kg b.w; i.p., twice a week for 30 days) generated oxidative stress in mice through accumulation of reactive oxygen species and increased lipid peroxidation, and levels of certain liver marker enzymes (ALT, AST, ALP) with decrease in levels of GSH and certain other antioxidant enzymes (SOD, CAT, GR) in liver. Treatment with nano-chelidonine for 30 days after CdCl2 intoxication significantly reduced oxidative stress and lipid peroxidation and restored levels of GSH, cholesterol, triglyceride and antioxidant enzymes, showing ameliorative changes in histopathology of liver. Expression pattern of certain inflammatory and apoptotic signal proteins also indicated better hepato-protective abilities of nano-chelidonine, making it a more suitable protective drug than chelidonine against cadmium toxicity in mice.

[In vitro transdermal delivery of the active fraction of xiangfusiwu decoction based on principal component analysis].

The objective of the present study was to establish a method based on principal component analysis (PCA) for the study of transdermal delivery of multiple components in Chinese medicine, and to choose the best penetration enhancers for the active fraction of Xiangfusiwu decoction (BW) with this method. Improved Franz diffusion cells with isolated rat abdomen skins were carried out to experiment on the transdermal delivery of six active components, including ferulic acid, paeoniflorin, albiflorin, protopine, tetrahydropalmatine and tetrahydrocolumbamine. The concentrations of these components were determined by LC-MS/MS, then the total factor scores of the concentrations at different times were calculated using PCA and were employed instead of the concentrations to compute the cumulative amounts and steady fluxes, the latter of which were considered as the indexes for optimizing penetration enhancers. The results showed that compared to the control group, the steady fluxes of the other groups increased significantly and furthermore, 4% azone with 1% propylene glycol manifested the best effect. The six components could penetrate through skin well under the action of penetration enhancers. The method established in this study has been proved to be suitable for the study of transdermal delivery of multiple components, and it provided a scientific basis for preparation research of Xiangfusiwu decoction and moreover, it could be a reference for Chinese medicine research.

Cytotoxicity and apoptotic signalling cascade induced by chelidonine-loaded PLGA nanoparticles in HepG2 cells in vitro and bioavailability of nano-chelidonine in mice in vivo.

Poor oral bioavailability of chelidonine, a bio-active ingredient of Chelidonium majus, showing anti-cancer potentials against cancer cells with multidrug resistance, makes its optimal use rather limited. To address this problem, we encapsulated chelidonine in biodegradable poly(lactide-co-glycolide) (PLGA) polymers and evaluated nano-chelidonine's (NCs) anti-cancer efficacy vis-à-vis free chelidonine (FC) against HepG2 cells and also evaluated its bioavailability in mice. Physicochemical characteristics indicated that stable spherical NC were formed in nanometer size range (123±1.15 nm) with good yield (86.34±1.91%), better encapsulation efficiency (82.6±0.574%), negative surface charge (-19.6±2.48 mV) and ability of prolonged and sustained release of chelidonine. Fourier transform infrared analysis revealed that NC resembled similar peaks as that of FC suggesting effective encapsulation in PLGA. NC exhibited rapid cellular uptake and stronger apoptotic effect (∼46.6% reduced IC50 value) than FC, blocking HepG2 cells at G2/M phase. p53, cyclin-D1, Bax, Bcl-2, cytochrome c, Apaf-1, caspase-9 and caspase-3 expressions also corroborated well to suggest greater anticancer potentials of NC. Our in vivo studies demonstrated NC to be more bio-available than FC and showed a better tissue distribution profile without inducing any toxicity (100 mg/kg bw) in mice. Unlike FC, NC could permeate into brain tissue, indicating thereby NC's better potentials for use in therapeutic oncology.

The pharmacokinetics of chelerythrine solution and chelerythrine liposomes after oral administration to rats.

Chelerythrine is a quaternary benzo[c]phenanthridine alkaloid which has many potent pharmacological effects and can dissolve well in water; dihydrochelerythrine has recently been identified as a chelerythrine metabolite in rat. Most methods of preparation of liposomes suffer from the drawback of poor incorporation of water-soluble drugs. The emulsion/solvent evaporation method is a relatively simple and efficient way to prepare liposomes loaded with hydrophilic drugs. The aim of this study was therefore to find a suitable formulation to enhance the incorporation of chelerythrine into liposomes by the emulsion/solvent evaporation method and so improve the therapeutic efficacy of chelerythrine. Results showed that the chelerythrine-liposome has been successfully prepared by the emulsion/solvent evaporation method: the entrapment efficiency of chelerythrine was higher at 78.6 %, and the drug loadings reached 7.8 %. The relative bioavailability of chelerythrine and its dihydro derivative in liposomes was significantly increased compared with that of the chelerythrine solution. The area under the plasma concentration-time curve values of chelerythrine and dihydrochelerythrine after oral administration of chelerythrine-liposomes were 4.83-fold and 2.02 higher than those obtained with the chelerythrine solution. The half time and peak concentrations of chelerythrine and dihydrochelerythrine were also higher for chelerythrine-liposomes than that for chelerythrine. In contrast, the total body clearance and apparent volume of distribution were lower for chelerythrine-liposomes in comparison to the respective parameters for the chelerythrine solution. It can thus be concluded that incorporation into liposomes prolonged chelerythrine retention within the systemic circulation.

Development and validation of an LC-ESI-MS/MS method for the determination of nitidine chloride in rat plasma.

A new liquid chromatography-electrospray ionization-mass/mass spectrometry (LC-ESI-MS/MS) assay method has been developed and validated for the quantification of nitidine chloride (NC), an anti-cancer bioactive substance of Zanthoxylum nitidum (Roxb.) DC. plants, in rat plasma using carbamazepine as an internal standard (I.S.). The NC and I.S. were extracted from rat plasma by acetonitrile protein procedure. Chromatographic separation was carried out with a C(18) column (2.1 mm × 150 mm, 3 µm) with a security guard C18 column (4 mm × 20 mm, 3 µm). The mobile phase consisted of acetonitrile-10 mM ammonium acetate buffer solution-formic acid (35:65:0.2, v/v/v) and delivered at the flow rate of 0.25 mL/min. LC-ESI-MS/MS was performed on a triple-quadrupole mass spectrometry equipped with electrospray ionization (ESI) and positive multiple reaction monitoring (MRM). Target ions were monitored at [M](+)m/z 348.2 for NC and [M]⁺ m/z 237.2 for I.S. The method was linear over the concentration range of 5.0-1500.0 ng/mL. The intra- and inter-day relative standard deviations of the assay were less than 5.0%. The lower limit of quantification was 5.0 ng/mL. The developed method was successfully applied to the estimation of the pharmacokinetic parameters of NC by intravenous administration to rats.

Effect of wine and vinegar processing of Rhizoma Corydalis on the tissue distribution of tetrahydropalmatine, protopine and dehydrocorydaline in rats.

Vinegar and wine processing of medicinal plants are two traditional pharmaceutical techniques which have been used for thousands of years in China. Tetrahydropalmatine (THP), dehydrocorydaline (DHC) and protopine are three major bioactive molecules in Rhizoma Corydalis. In this study, a simple and reliable HPLC method was developed for simultaneous analysis of THP, DHC and protopine in rat tissues after gastric gavage administration of Rhizoma Corydalis. The validated HPLC method was successfully applied to investigate the effect of wine and vinegar processing on the compounds' distribution in rat tissues. Our results showed that processing mainly affect the T(max) and mean residence time (MRT) of the molecules without changing their C(max) and AUC(0-24)( )(h) Vinegar processing significantly increased the T(max) of DHC in heart, kidney, cerebrum, cerebrellum, brain stem and striatum and prolonged the T(max) of protopine in brain. No significant changes were observed on the T(max) of THP in rat tissues after vinegar processing. Wine processing reduced the T(max) of protopine and DHC in liver and spleen and T(max) of protopine in lung, but increased the T(max) of THP in all the rat tissues examined. To our knowledge, this is the first report on the effects of processing on the tissue distribution of the bioactive molecules from Rhizoma Corydalis.

In vitro and in vivo evaluation of sanguinarine liposomes prepared by a remote loading method with three different ammonium salts.

Sanguinarine liposomes were prepared by a remote loading method using three different ammonium salts. A series of studies, including in vitro release, in vitro and in vivo anti-tumor effects and pharmacokinetics in rats, were conducted. The three liposomes showed pH-sensitive release characteristics in vitro, but there were obvious variations in their release profiles. Among the three liposomes, the liposomes made using ammonium citrate and phosphate possessed better anti-tumor activity in vitro and in vivo, compared with the liposome using ammonium sulfate. Pharmacokinetics test results in rats indicated that sanguinarine liposomes have notably elevated AUC (P<0.05) and markedly lower CL (P<0.05) compared with the solution, but there were no obvious differences between the three liposomes. The present study may be useful for better understanding and better choice of a suitable ammonium salt for the remote loading method.

Identification of benzo[c]phenanthridine metabolites in human hepatocytes by liquid chromatography with electrospray ion-trap and quadrupole time-of-flight mass spectrometry.

The metabolism of the benzo[c]phenanthridine alkaloids was studied using human hepatocytes which are an excellent model system for biotransformation studies. For analysis of the alkaloids and their metabolites, an electrospray quadrupole ion-trap mass spectrometry (ESI ion-trap MS) connected to a reversed phase chromatographic system based on cyanopropyl modified silica was used. The optimized experimental protocol allowed simultaneous analysis of the alkaloids and their metabolites and enabled study of their uptake into and interconversion in human hepatocytes. The results show that formation of the dihydro metabolite which may be followed by specific O-demethylenation/O-demethylation processes, is probably the main route of biotransformation (detoxification) of the benzo[c]phenanthridines in human hepatocytes. The structure of the main O-demethyl metabolite (2-methoxy-12-methyl-12,13-dihydro-[1,3]dioxolo[4',5':4,5]benzo[1,2-c]phenanthridin-1-ol; 336.1 m/z,) was proposed by the multi-stage MS and quadrupole time-of-flight MS methods using chemically synthesized standard.