Enhanced anti-amnestic effect of donepezil by Ginkgo biloba extract (EGb 761) via further improvement in pro-cholinergic and antioxidative activities.

ETHNOPHARMACOLOGICAL RELEVANCE: EGb 761 is a standardized dry extract of Ginkgo biloba L. leaves traditionally used by Eastern Asia and has been associated with beneficial effects on neurodegeneration disorders, including Alzheimer's disease. AIM OF THE STUDY: Since beneficial interactions between EGb 761 and donepezil have been observed in previous clinical studies, the current study was proposed aiming to further explore related mechanisms from both pharmacokinetics and pharmacodynamics aspects. MATERIALS AND METHODS: Pharmacodynamic interactions were studied in scopolamine-induced cognitive impairment rats received two-weeks treatment of vehicle, EGb 761 and/or donepezil by the Morris water maze test and ex vivo evaluation of biomarkers of cholinergic transmission and oxidative stress in rat brain. In the meantime, pharmacokinetic profiles of donepezil and bilobalide were obtained and compared among all treatment groups. In addition, impact of the bioavailable EGb 761 components on donepezil brain penetration was evaluated with the hCMEC/D3 cell monolayer model. RESULTS: Scopolamine-induced rats with co-treatment of EGb 761 and donepezil had significantly improved cognitive function in the Morris water maze test with increased brain levels of superoxide dismutase and decreased brain levels of acetylcholinesterase and malondialdehyde than that with treatment of only EGb 761 or donepezil. Despite such beneficial pharmacodynamics outcomes, the two-week co-treatment of EGb 761 and donepezil did not alter the plasma pharmacokinetics and brain uptake of donepezil or bilobalide, which was further verified in the hCMEC/D3 monolayer model. CONCLUSION: Co-administration of EGb 761 and donepezil exerted better anti-amnestic effect via further enhanced pro-cholinergic and antioxidative effects of EGb 761 or donepezil in scopolamine-induced cognitive impairment rat without alteration in their systemic/brain exposure.

Pure omega 3 polyunsaturated fatty acids (EPA, DPA or DHA) are associated with increased plasma levels of 3-carboxy-4-methyl-5-propyl-2-furanpropanoic acid (CMPF) in a short-term study in women.

3-Carboxy-4-methyl-5-propyl-2-furanpropanoic acid (CMPF) is a metabolite of furan fatty acids found in plasma and urine of humans after consumption of foods containing these fatty acids. Recently, CMPF has been identified as a prominent metabolite following the consumption of either fish oil, fish oil fatty acid-ethyl esters or diets rich in fish. As furan fatty acids are known to occur in fish and fish oils (at a low level), it is possible that in these studies the CMPF in plasma originated from furan fatty acids. We report the plasma CMPF levels in 10 healthy women who consumed 1 gram of pure eicosapentaenoic acid (EPA), or docosapentaenoic acid (DPA) or docosahexaenoic acid (DHA), or olive oil daily for 6 days, in a cross-over study. The supplemented omega 3 fatty acids contained no detectable levels of furan fatty acids. The plasma CMPF and omega 3 fatty acid levels were measured by LC-MS/MS. Consumption of pure omega 3 fatty acids led to a significant increase in the plasma CMPF levels, but not with olive oil (from 1.6 to 2.5-fold compared with baseline). The plasma free fatty acid levels of EPA, DPA and DHA also increased significantly when they were supplemented (p < 0.05). Significant positive correlations existed between the plasma free fatty acid DPA and DHA levels (p < 0.05 and r = +0.49 to +0.81), but not between the EPA and CMPF levels. These data suggest that purified long chain omega 3 fatty acids may be precursors of CMPF; however the metabolic pathway(s) from omega 3 fatty acids to CMPF remain to be elucidated.

Influence of organic anion transporter 1/3 on the pharmacokinetics and renal excretion of ginkgolides and bilobalide.

ETHNOPHARMACOLOGICAL RELEVANCE: The major terpene lactones of ginkgo biloba extract (GBE) include ginkgolide A, B, C and bilobalide are used for the protection of cardiovascular, cerebrovascular and neurodegenerative diseases. Terpene lactones are orally bioavailable and predominantly eliminated via the renal pathway. However, information on the transporters involved in the pharmacokinetics (PK) and renal excretion of terpene lactones is limited. AIM OF THE STUDY: The objective of this study is to assess the role of OAT1/3 which are important transporters in the human kidney in the PK and renal excretion ginkgolide A, B, C and bilobalide. MATERIALS AND METHODS: Uptake of ginkgolide A, B, C and bilobalide in Madin-Darby Canine Kidney (MDCK) and human embryonic kidney 293 (HEK293) cells overexpressing OAT1 or OAT3, respectively were studied. To verify the result from in vitro cell models, the studies on PK, kidney accumulation and urinary excretion of ginkgolide A, B, C and bilobalide were carried out in rats. RESULTS: The result showed that ginkgolide A, B, C and bilobalide are low-affinity substrates of OAT1/3. Following co-administration with probenecid, a typical inhibitor of OAT1/3, the rat plasma concentrations of ginkgolide A, B, C and bilobalide increased significantly. AUC showed a significant increase in the probenecid-treated rats compared to control rats (893.48 vs. 1123.85, 314.91 vs. 505.74, and 2724.97 vs. 3096.40 µg/L*h for ginkgolide A, B and bilobalide, respectively), while the clearance of these compounds significantly decreased. The accumulation of ginkgolide A, B and bilobalide in the kidney of the probenecid-treated rats was reduced by 1.8, 2.4, and 1.5-fold, respectively; further reducing the cumulative urinary recovery of these compounds. CONCLUSION: The findings indicated that ginkgolide A, B and bilobalide are excreted via OAT1/3-mediated transport in the kidney and OAT1/3 inhibitor significantly influence the PK ginkgolides and bilobalide.

Novel PAradigm to improve Inflammatory burden in end stage Renal disease (rePAIR): study protocol for a randomized controlled trial.

BACKGROUND: Given the importance of inflammation as a predictor of poor outcomes in End Stage Renal Disease (ESRD), reductions in inflammatory biomarkers have been proposed as a critical target in this population. This study targets chronic periodontitis, an oral inflammatory disease of microbial etiology causing persistent inflammation in ESRD. Unlike the previously reported episodic periodontal interventions, we propose to control periodontal inflammation with a continuous maintenance and oral health behavior modifications. We hypothesize that this strategy will improve systemic inflammation and oxidative stress, oral health and quality of life within the 6-month observation period. METHODS: The rePAIR (novel PAradigm to improve Inflammatory burden in ESRD) study is a pilot and feasibility, parallel-arm, and randomized controlled clinical trial that will recruit 72 ESRD subjects with periodontitis in a model of computerized block randomization. This trial aims to compare the effect of standard-of-care vs. repeated non-surgical periodontal therapy on systemic and oral inflammatory burden. This trial will recruit ESRD adult patients with periodontitis older than 21 years old with a minimum of 12 teeth and no history of periodontal treatment within a year. The trial will examine serum C-reactive protein (CRP) (primary outcome) as a biomarker of inflammation as well as interleukin-6 (IL-6), F2 isofurans and F2 isoprostanes (secondary outcomes) and compare their difference between groups from baseline to 6 months. The trial will also compare the difference between groups in patient-centered and clinical oral outcomes from baseline to 6 months. DISCUSSION: The trial follows a rigorous and transparent study design capturing elements such as pre-specified eligibility criteria, pre-specified primary and secondary outcomes, detailed intervention description to allow replication, intervention random allocation and concealment, blinding in outcome assessment, appropriate sample size calculations, explanation of interim analysis, as per CONSORT Guidelines. Further, gender diversity is secured not only at recruitment but also throughout the trial and during the analysis. Therefore, treatment response outcomes will be examined per gender category. In order to manage anticipated problems, the protocol has included alternative approaches. TRIAL REGISTRATION: ClinicalTrials.gov, NCT03241511 . Registered on 7 August 2017.

Metabolomic markers of fatigue: Association between circulating metabolome and fatigue in women with chronic widespread pain.

BACKGROUND: Fatigue is a sensation of unbearable tiredness that frequently accompanies chronic widespread musculoskeletal pain (CWP) and inflammatory joint disease. Its mechanisms are poorly understood and there is a lack of effective biomarkers for diagnosis and onset prediction. We studied the circulating metabolome in a population sample characterised for CWP to identify biomarkers showing specificity for fatigue. MATERIAL AND METHODS: Untargeted metabolomic profiling was conducted on fasting plasma and serum samples of 1106 females with and without CWP from the TwinsUK cohort. Linear mixed-effects models accounting for covariates were used to determine relationships between fatigue and metabolites. Receiver operating curve (ROC)-analysis was used to determine predictive value of metabolites for fatigue. RESULTS: While no association between fatigue and metabolites was identified in twins without CWP (n=711), in participants with CWP (n=395), levels of eicosapentaenoate (EPA) ω-3 fatty acid were significantly reduced in those with fatigue (ß=-0.452±0.116; p=1.2×10-4). A significant association between fatigue and two other metabolites also emerged when BMI was excluded from the model: 3-carboxy-4-methyl-5-propyl-2-furanpropanoate (CMPF), and C-glycosyltryptophan (p=1.5×10-4 and p=3.1×10-4, respectively). ROC analysis has identified a combination of 15 circulating metabolites with good predictive potential for fatigue in CWP (AUC=75%; 95% CI 69-80%). CONCLUSION: The results of this agnostic metabolomics screening show that fatigue is metabolically distinct from CWP, and is associated with a decrease in circulating levels of EPA. Our panel of circulating metabolites provides the starting point for a diagnostic test for fatigue in CWP.

Hyperbaric oxygen therapy is not associated with oxidative stress assessed using plasma F2-isoprostanes and isofurans.

BACKGROUND AND AIMS: Hyperbaric oxygen (HBO) therapy is increasingly used in medical practice as a means of enhancing the formation of collagen matrix and angiogenesis, thus promoting healing in wounds and necrotic tissue. However, there are concerns that oxygen can also associate with increased production of oxygen free radicals and oxidative stress. F2-Isoprostanes (F2-IsoPs) formed by non-enzymatic oxidation of arachidonic acid (AA) are reliable measures for assessing oxidative stress in vivo. In addition, under conditions of high oxygen tension isofurans (IsoFs) are preferentially formed from AA and are considered to better reflect oxidative stress in the setting of high oxygen tension. This study aimed to measure plasma IsoFs and F2-IsoP in patients receiving HBO therapy to treat osteonecrosis secondary to radiation therapy. Our hypothesis was that IsoFs would continue to rise with increasing oxygen pressures in contrast to F2-IsoPs whose synthesis would be reduced. METHODS: Twelve patients receiving hyperbaric therapy to treat osteonecrosis secondary to radiation therapy were studied during hyperbaric treatment. Blood samples were collected prior to, during and after cessation of HBO therapy that lasted for 119min. Seven serial blood samples were collected for measurement of plasma F2-IsoPs and IsoFs, blood gases and haemoglobin. RESULTS: Oxygen saturation and venous oxygen partial pressure (PvO2) rose significantly during hyperbaric therapy. However, there were no significant changes in plasma IsoFs or F2-IsoPs during the hyperbaric therapy session. CONCLUSION: In this study of patients with osteonecrosis, HBO therapy at a maximum pressure of 2.4atm with up to 100% oxygen did not worsen oxidative stress assessed using plasma F2- IsoFs and IsoPs.

UHPLC-ESI-MS/MS determination and pharmacokinetics of pinoresinol glucoside and chlorogenic acid in rat plasma after oral administration of Eucommia ulmoides Oliv extract.

This study aimed to develop a specific UHPLC-ESI-MS/MS method for simultaneous determination and pharmacokinetics of pinoresinol glucoside and chlorogenic acid in rat plasma after oral administration of Eucommia ulmoides. The chromatographic separation was achieved on a Hypersil GOLD column with gradient elution by using a mixture of 0.1% formic acid aqueous solution and acetonitrile as the mobile phase at a flow rate of 200 µL/min. A tandem mass spectrometric detection was conducted using multiple-reaction monitoring via an electrospray ionization source in negative ionization mode. Samples were pre-treated by a single-step protein precipitation with acetonitrile, and bergenin was used as internal standard. After oral administration of 3 mL/kg E. ulmoides extract in rats, the maximum plasma concentrations of pinoresinol glucoside and chlorogenic acid were 57.44 and 61.04 ng/mL, respectively. The times to reach the maximum plasma concentration were 40.00 and 23.33 min for pinoresinol glucoside and chlorogenic acid, respectively. The intra- and inter-day precision (RSD) values for the two analytes were <2.46 and 5.15%, respectively, and the accuracy (RE) values ranged from -12.76 to 0.00. This is the first study on pharmacokinetics of bioactive compounds in rat plasma after oral administration of E. ulmoides extract.

Simultaneous Determination and Pharmacokinetic Study of Six Components in Rat Plasma by HPLC-MS/MS after Oral Administration of Acanthopanax sessiliflorus Fruit Extract.

A specific and reliable HPLC-MS/MS method was developed and validated for the simultaneous determination of protocatechuic acid (PCA), scopolin, (-)-pinoresinol-4,4'-di-O-ß-d-glucopyranoside (PDG), acanthoside D, acanthoside B and hyperin in rat plasma for the first time. The analytes were separated on a C18 column (50 × 2.1 mm, 1.8 µm) and a triple-quadrupole mass spectrometer equipped with an electrospray ionization (ESI) source was used for detection. The rat plasma sample was prepared using the protein precipitation procedure. The calibration curves were linear over a concentration range of 1.2-1200.0 ng/mL for PCA, 0.96-960.0 ng/mL for scopolin, 1.12-1120.0 ng/mL for PDG, 1.32-1320.0 ng/mL for acanthoside D, 0.99-990.0 ng/mL for acanthoside B and 1.01-1010.0 ng/mL for hyperin. The intra-day and inter-day precision was less than 11.4% and the relative error (RE) was all within ±15%. The validated method was successfully applied to assess the pharmacokinetics characteristics after the extracts of Acanthopanax sessiliflorus fruits were orally administered to the Sprague-Dawley rat.

Metabolomics analysis of serum 25-hydroxy-vitamin D in the Alpha-Tocopherol, Beta-Carotene Cancer Prevention (ATBC) Study.

BACKGROUND: Vitamin D has been discussed in the context of cardiovascular disease, cancer, bone health and other outcomes. Epidemiological studies have reported on the importance of vitamin D in cancer prevention and treatment. The discovery of vitamin D-associated metabolites through agnostic metabolomics analyses offers a new approach for elucidating disease aetiology and health-related pathway identification. METHODS: Baseline serum 25-hydroxy-vitamin D [25(OH)D] and 940 serum metabolites were measured in 392 men from eight nested cancer case-control studies in the Alpha-Tocopherol, Beta-Carotene Cancer Prevention Study of Finnish male smokers (aged 50-69 years). The metabolomic profiling was conducted using mass spectrometry. We used linear regression to estimate the standardized beta-coefficient as the effect metric for the associations between metabolites and 25(OH)D levels. RESULTS: A majority of the metabolites associated with 25(OH)D were of lipid origin, including 3-carboxy-4-methyl-5-propyl-2-furanpropanoic acid (CMPF) [beta-estimate 0.38 per 1 standard deviation (SD) increment], stearoyl-arachidonoyl-glycerophosphoethanolamine (GPPE) (-0.38 per SD) and two essential fatty acids: eicosapentaenoate (EPA; 0.17 per SD) and docosahexaenoate (DHA; 0.13 per SD). Each of these lipid metabolites was associated with 25(OH)D at the principal components corrected P-value of 3.09 × 10-4 CONCLUSIONS: The large number of metabolites, particularly lipid compounds, found to be associated with serum 25(OH)D provide new biological clues relevant to the role of vitamin D status and human health outcomes. The present findings should be re-examined in other metabolomics studies of diverse populations.

Determination of Ginkgolides A, B, C, J and Bilobalide in Plasma by LC-ESI (-)/MS/MS (QQQ) and its Application to the Pharmacokinetic Study of Ginkgo Biloba Extract in Rats.

A simple, rapid, and specific high-performance liquid chromatograph coupled with a tandem mass spectrometry method has been developed and validated for the quantification of ginkgolides in rat plasma, and the main pharmacokinetic parameters of ginkgolides after oral administration of Ginkgo biloba extract (GBE) was acquired. Methods: Plasma samples were pretreated with ethyl acetate extraction. Sulfamethoxazole was used as the internal standard (IS). Chromatographic separation was achieved on an Eclipse XDB-C18 column (2.1 mm×150 mm, 5 µm) with a mobile phase consisting of methanol/0.1% formic acid water (gradient elution: 0~25 min (77:23)→(60:40), V/V) at a flow rate of 0.3 mL·min-1. The detection was performed on a triple quadruple tandem mass spectrometer using an electrospray ionization (ESI) source for 25 min. The detection was operated by multiple reaction monitoring(MRM) under negative ionization mode of the transitions of m/z 325→163 for BB, 469→423 for GJ, 439→125 for GC, 453→351 for GA, 423→367 for GB and of m/z 252→156 for sulfamethoxazole (IS) respectively. Results: The pharmacokinetic properties of BB, GJ, GA, GB and GC were in line with the open 2-compartment model after oral administration of GBE in rats; The pharmacokinetic parameters of various lactones were calculated, and drugs-time curve and the curve fitting diagram of 5 ginkgolides were drew; The absorption and distribution rate of BB, GJ, GA, GB and GC were fast in rats in vivo, and half-life of absorption was less than 3 h. Conclusion: The developed LC-ESI (-)/MS/MS (QQQ) method was successfully applied to assess the pharmacokinetic parameters and oral bioavailability of ginkgolides in rats after administration of GBE, which can provide basis for further clinical efficacy studies.

Serum metabolomics profiles in response to n-3 fatty acids in Chinese patients with type 2 diabetes: a double-blind randomised controlled trial.

We aimed to investigate the change of serum metabolomics in response to n-3 fatty acid supplements in Chinese patients with type 2 diabetes (T2D). In a double-blind parallel randomised controlled trial, 59 Chinese T2D patients were randomised to receive either fish oil (FO), flaxseed oil (FSO) or corn oil capsules (CO, served as a control group) and followed up for 180 days. An additional 17 healthy non-T2D participants were recruited at baseline for cross-sectional comparison between cases and non-cases. A total of 296 serum metabolites were measured among healthy controls and T2D patients before and after the intervention. Serum 3-carboxy-4-methyl-5-propyl-2-furanpropanoate (CMPF) (P-interaction = 1.8 × 10(-7)) was the most significant metabolite identified by repeated-measures ANOVA, followed by eicosapentaenoate (P-interaction = 4.6 × 10(-6)), 1-eicosapentaenoylglycerophosphocholine (P-interaction = 3.4 × 10(-4)), docosahexaenoate (P-interaction = 0.001), linolenate (n-3 or n-6, P-interaction = 0.005) and docosapentaenoate (n-3, P-interaction = 0.021). CMPF level was lower in T2D patients than in the healthy controls (P = 0.014) and it was significantly increased in the FO compared with CO group (P = 1.17 × 10(-7)). Furthermore, change of CMPF during the intervention was negatively correlated with change of serum triglycerides (P = 0.016). In conclusion, furan fatty acid metabolite CMPF was the strongest biomarker of fish oil intake. The association of CMPF with metabolic markers warrants further investigation.

Enhancement of site specific delivery of diloxanide furoate as an antiamoebic drug.

The basic aim of the present research work is to deliver the diloxanide furoate (DF) at specific area using pectin microspheres. The microspheres were prepared by spray drying method and cross-linked by zinc acetate. Different concentrations of polymer (pectin 0.5-3%) and cross-linking agent (0-3% w/v in a mixture of ethanol:water) are taken to optimize the entrapment efficiency, swelling behavior, size and first 6h in-vitro release in simulated gastric fluids. Optimized formulation was characterized in the terms of in-vitro release, in-vivo drug disposition in various organs and in the blood of Sprague-Dawley albino rats and in-vivo gastrointestinal tract transit behavior using X-ray imaging method on albino rabbits. Findings suggested that microspheres containing a concentration of polymer (2% w/v) have average size of 100-500 µm, entrapment efficiency 85.82 ± 0.5 with swelling index 18.77 ± 5.21. In-vitro results and in-vivo gastric transit behavior (using X-ray imaging) have shown no release in first 3-6h that proved the colon specific delivery of DF. The results also suggested that the above approach have not only site specific delivery, but it improves the conversion of active drug by increasing the enzyme mediated hydrolytic degradation of DF due to the presence of polysaccharide polymer:water gel complex.

A UPLC-MS Method for Simultaneous Determination of Geniposidic Acid, Two Lignans and Phenolics in Rat Plasma and its Application to Pharmacokinetic Studies of Eucommia ulmoides Extract in Rats.

The bark of Eucommia ulmoides is a well-known Chinese herbal medicine that is used to regulate blood pressure and reduce blood sugar and fats, as well as an antioxidant and antimicrobial agent. Here we describe the development of a sensitive ultrahigh performance liquid chromatography-tandem mass spectrum method for the simultaneous determination of five major active ingredients of E. ulmoides bark extract, namely, geniposidic acid (GA), protocatechuic acid (PCA), chlorogenic acid (CA), (+)-pinoresinol di-O-ß-D-glucopyranoside (PDG) and (+)-pinoresinol 4'-O-ß-D-glucopyranoside (PG), in rat plasma. The preliminary steps in the plasma analysis were the addition of an internal standard and acidification (0.1 % formic acid), followed by protein precipitation with methanol. Separation of the active ingredients was performed on an ACQUITY UPLC® BEH C18 column (2.1 × 50 mm; internal diameter 1.7 µm) at a flow rate of 0.35 mL/min, with acetonitrile/water containing 0.1 % formic acid as the mobile phase. Detection was performed on a triple quadrupole tandem mass spectrometer via electrospray ionization source with positive and negative ionization modes. All calibration curves showed good linearity (r ≥ 0.997) over the concentration range with the low limit of quantification between 4.45 and 54.9 ng/mL. Precision was evaluated by intra- and inter-day assays, and the percentages of the relative standard deviation were all within 15 %. Extraction efficiency and matrix effect were 84.3-102.4 % and 98.1-112.2 %, respectively. The validated method was successfully applied to the pharmacokinetic study in rats after oral administration of E. ulmoides extract. The results indicate that the pharmacokinetic properties of GA differ from those of PCA, CA, PDG and PG, respectively.

Method development for quantification of the environmental neurotoxin annonacin in Rat plasma by UPLC-MS/MS and application to a pharmacokinetic study.

Annonacin is an environmental neurotoxin identified in the pulp of several fruits of the Annonaceae family (for example in Annona muricata, Asimina triloba), whose consumption was linked with the occurrence of sporadic atypical Parkinsonism with dementia. Pharmacokinetic parameters of this molecule are unknown. A method for its quantification in Rat plasma was developed, using its analogue annonacinone as an internal standard. Extraction from plasma was performed using ethylacetate with a good recovery. Quantification was performed by UPLC-MS/MS in SRM mode, based on the loss of the γ-methyl-γ-lactone (-112amu) from the sodium-cationized species [M+Na](+) of both annonacin and internal standard. The limit of quantification was 0.25ng/mL. Despite strong matrix effects, a good linearity was obtained over two distinct ranges 0.25-10ng/mL and 10-100ng/mL. The intra- and inter-day precisions (RSD) were lower than 10%, while accuracy was within ±10%. This method was applied to a pharmacokinetic study in the Rat. After oral administration of 10mg/kg annonacin, a Cmax of 7.9±1.5ng/mL was reached at Tmax 0.25h; T1/2 was 4.8±0.7h and apparent distribution volume was 387.9±64.6L. The bioavailability of annonacin was estimated to be 3.2±0.3% of the ingested dose.

CMPF does not associate with impaired glucose metabolism in individuals with features of metabolic syndrome.

OBJECTIVE: 3-carboxy-4-methyl-5-propyl-2-furanpropanoic acid (CMPF) is a metabolite produced endogenously from dietary sources of furan fatty acids. The richest source of furan fatty acids in human diet is fish. CMPF was recently shown to be elevated in fasting plasma in individuals with gestational diabetes and type 2 diabetes, and mechanistically high level of CMPF was linked to ß cell dysfunction. Here we aimed to study the association between plasma CMPF level and glucose metabolism in persons with impaired glucose metabolism. METHODS: Plasma CMPF concentration was measured from plasma samples of the study participants in an earlier controlled dietary intervention. All of them had impaired glucose metabolism and two other characteristics of the metabolic syndrome. Altogether 106 men and women were randomized into three groups for 12 weeks with different fish consumption (either three fatty fish meals per week, habitual fish consumption or maximum of one fish meal per week). Associations between concentration of CMPF and various glucose metabolism parameters at an oral glucose tolerance test at baseline and at the end of the study were studied. RESULTS: Fasting plasma CMPF concentration was significantly increased after a 12-week consumption of fatty fish three times per week, but the concentration remained much lower compared to concentrations reported in diabetic patients. Increases of plasma CMPF concentrations mostly due to increased fish consumption were not associated with impaired glucose metabolism in this study. Instead, elevated plasma CMPF concentration was associated with decreased 2-hour insulin concentration in OGTT. CONCLUSIONS: Moderately elevated concentration of CMPF in plasma resulting from increased intake of fish is not harmful to glucose metabolism. Further studies are needed to fully explore the role of CMPF in the pathogenesis of impaired glucose metabolism. TRIAL REGISTRATION: ClinicalTrials.gov NCT00573781.

Arctigenic acid, the key substance responsible for the hypoglycemic activity of Fructus Arctii.

We have reported the antidiabetic activity of the total lignans from Fructus arctii (TLFA) against alloxan-induced diabetes in mice and rats. In this study, arctigenic acid was found to be the main metabolite in rat plasma detected by UPLC/MS and HPLC/MS/MS after oral administration of TLFA. For the first time, its hypoglycemic activity and acute oral toxicity were evaluated in Goto-Kakizaki (GK) rats, a spontaneous type 2 diabetic animal model, and ICR mice respectively. GK rats were orally given arctigenic acid (50 mg/kg) twice daily before each meal for 12 weeks. The treatment reduced the elevated plasma glucose, glycosylated hemoglobin and showed significant improvement in glucose tolerance in glucose fed hyperglycemic GK rats. We found that the hypoglycemic effect of arctigenic acid was partly due to the stimulation on insulin secretion, whereas the body weight was not affected by arctigenic acid administration in GK rats. Meanwhile, there was no observable acute toxicity of arctigenic acid treatment at the dosage of 280 mg/kg body weight daily in the acute 14-day toxicity study in mice. This study demonstrates that arctigenic acid may be the main metabolite in the rat serum after oral administration of TLFA, which showed significant hypoglycemic effect in GK rats, and low acute toxicity in ICR mice. The result prompts us that arctigenic acid is the key substance responsible for Fructus Arctii antidiabetic activity and it has a great potential to be further developed as a novel therapeutic agent for diabetes in humans.

Simultaneous quantification of curdione, furanodiene and germacrone in rabbit plasma using liquid chromatography-tandem mass spectrometry and its application to a pharmacokinetic study.

A simple, rapid and sensitive method was developed for the simultaneous quantification of curdione, furanodiene and germacrone in rabbit plasma using a LC-MS/MS analysis. The plasma sample preparation was a simple deproteinization by the addition of 3 vols of acetonitrile followed by centrifugation. The analytes and internal standard (IS) costunolide were separated on a Zorbax SB-C18 column (3.5 µm, 2.1 × 100 mm) with mobile phase of methanol-water (90:10, v/v) containing 0.1% formic acid at a flow rate of 0.3 mL/min with an operating temperature of 25°C. Detection was carried out by atmospheric pressure chemical ionization in positive ion selected reaction monitoring mode. Linear detection responses were obtained for the three test compounds ranging from 5 to 5000 ng/mL and the lower limits of quantitation were 5-10 ng/mL. The intra- and inter-day precisions (relative standard deviations) were within 9.4% for all analytes, while the deviation of assay accuracies was within ±10.0%. The average recoveries of analytes were >80.0%. All analytes were proved to be stable during all sample storage, preparation and analytical procedures. The method was successfully applied to the pharmacokinetic study of the three compounds after vaginal drug delivery of Baofukang suppository to rabbit.

Determination of xanthatin by ultra high performance liquid chromatography coupled with triple quadrupole mass spectrometry: application to pharmacokinetic study of xanthatin in rat plasma.

A sensitive, specific and rapid ultra high performance liquid chromatography tandem mass spectrometry (UHPLC-MS/MS) method has been established to study pharmacokinetic properties of xanthatin. Xanthatin, a compound which belongs to sesquiterpene lactone group, was determined in rat plasma with psoralen as internal standard. Chromatographic separation was performed on an Agilent Zorbax Eclipse plus C18 column (50 mm × 2.1 mm, 3.5 µm) with gradient elution system at a flow rate of 0.3 mL/min. The mobile phase was composed of methanol and 0.1% formic acid water solution. Analysis was performed under a triple-quadruple tandem mass-spectrometer with an electrospray ionization (ESI) source via the multiple reaction monitoring (MRM) mode to determine xanthatin at [M+H](+)m/z 247.3→m/z 205.2 and that of IS at [M+H](+)m/z 187.1→m/z 143.0 within 5 min. The assay method exhibited good separation of xanthatin from the interference of endogenous substances. The lower limit of quantification (LLOQ) was 1 ng/mL, with a good linearity within the concentration range of 1-5000 ng/mL (r=0.9990). Intra-day and inter-day precision RSD was less than 9.27%; intra-day and inter-day accuracy was 88.48% and 102.25% respectively. The extraction recoveries of xanthatin range from 82.12% to 89.55%, and the extraction RSD was less than 9.01%. The established LC-ESI-MS/MS method is rapid and sensitive, which has been successfully applied to quantify xanthatin in rat plasma for the first time.

New insights regarding tissue Se and Hg interactions on oxidative stress from plasma IsoP and IsoF measures in the Canadian Inuit population.

Despite animal and in vitro studies demonstrating pro-oxidative effects of Hg, previous human work showed no relationship between tissue Hg and plasma levels of F2-isoprostanes (IsoPs), a whole-body oxidative stress marker. We hypothesized that another IsoP species, isofurans (IsoFs), was a more sensitive indicator of Hg-mediated oxidative stress, which can be modified by tissue Se status. A cross-sectional study was carried out involving individuals from a random subset (n = 233) of Inuit adults from a population-based survey (n = 2,595) of 36 Canadian Arctic Inuit communities to assess the relationships of plasma IsoPs to Se and Hg status indicators. F2-IsoPs were inversely correlated with blood Se (r = -0.186, P = 0.005) and toenail Se (r = -0.146, P = 0.044), but not correlated with Hg. IsoFs were inversely correlated with blood Se (r = -0.164, P = 0.014) and positively correlated with Hg (r = 0.228, P < 0.001) and Hg:Se (r = 0.340, P < 0.001). The strength of the correlations remained unchanged after multivariate adjustments. Multivariate analysis showed that F2-IsoPs were not positively associated with Hg but with Hg:Se (ß = 0.148, P = 0.021). We conclude that Se and Hg status and their interactions are important factors modulating F2-IsoP and IsoF levels such that the Inuit may be protected from Hg-induced oxidative stress because of their high Se status.

Plasma pharmacokinetics and tissue distribution of arctiin and its main metabolite in rats by HPLC-UV and LC-MS.

The pharmacokinetic profile of arctiin, the major active lignan in fruits of Arctium lappa L., was investigated. Its main meta"bolite arctigenin was identified by an LC-MS method, and an HPLC-UV technique was developed for the simultaneous quantification of the metabolite and arctiin in plasma and organs. Chromatographic separation was performed on an Agilent™ C18 HPLC column with acetonitrile and water by linear gradient elution. Arctiin and arctigenin were identified on-line by LC-MS. The pharmacokinetics and tissue distribution of arctiin and arctigenin were determined for the first time by using a simple, selective, and accurate HPLC method. The AUC0-t values of arctigenin were larger compared with arctiin after oral administration of arctiin. The concentration of the metabolite was significantly higher than the concentration of arctiin in the stomach and small intestine in rats after oral administration of arctiin, indicating that the stomach and small intestine were the major organs of arctiin metabolism. These findings could provide support for the clinical studies conducted with Fructus Arctii.