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A sample of 10 mM flurbiprofen in methanol (or ethanol) was photoirradiated with sixteen 8 W low-pressure quartz mercury lamps irradiated at 306 nm in a Panchum PR-2000 photochemical reactor. In total, four major photoproducts derived from each sample were observed from the HPLC chromatogram. The photoproducts were separated and their structures elucidated by various spectroscopic methods. Alternatively, using GC-MS, 11 major photoproducts were observed. A reaction scheme of flurbiprofen in methanol is proposed: the photochemical reaction routes occur mainly via esterification and decarboxylation, followed by oxidation with singlet oxygen to produce a ketone, alcohols and other derivatives., ((c) 2006 John Wiley & Sons, Ltd.)

Most human pyelonephritogenic Escherichia coli express the PapG II adhesin. However, the role of the PapG II adhesin in enhancing the establishment and persistence of E. coli infection in the kidney is controversial. A pyelonephritogenic strain, EC114, which possesses one copy of the papG II gene, but without other virulence factors (such as S/F1C-fimbriae, hemolysin, and cytotoxic necrotizing factor 1) was selected for the construction of a papG II mutant. The resulting papG II mutant was confirmed by polymerase chain reaction, Southern hybridization, and agglutination assay, and designated as MEC114. We compared MEC114 with the parental strain (EC114) for colonization ability in the bladder and kidney of female BALB/c mice, which were challenged transurethrally with 50 microl of a low (5x10(4) CFU (colony-forming unit)) or high (5x10(8) CFU) dose of EC114 or MEC114 and assessed 1, 3, and 7 days after inoculation. Geometric means of quantitative bacterial counts in the kidney were significantly decreased when challenged with MEC114 on day 3 after inoculation, at both low and high dose (P<0.05), as compared with EC114. On the seventh day, both strains were mainly cleared from the kidney. Renal biopsy showed a similar degree of inflammatory response to both strains 1, 3, and 7 days after inoculation. In brief, the PapG II adhesin can enhance the early establishment of E. coli infection in the kidney, but the bacteria do not maintain infection owing to the host immune response.

A rapid, accurate and reliable reversed-phase high-performance liquid chromatographic (HPLC) method for the determination of naproxen and its photodegradation products in methanol was developed and validated. An Inertsil 5-ODS-3V column (5 microm, C18, 250 x 4.6 mm i.d.) was used with a mobile phase of acetonitrile-methanol-1% HOAc in H2O (40:20:40, v/v/v). UV detection was set at 230 nm. The developed method satisfies system suitability criteria, peak integrity and resolution for the parent drug and its photoproducts. The intraday and interday standard deviations of five replicate determinations for five consecutive days at the working concentrations of 5.0, 10, 25, 50, and 100 microm were 0.23-0.98 with coefficients of variance (CVs) of between 0.96 and 4.56% for the former, and 0.14-1.15 with CVs of between 1.13 and 3.82% for the latter. The percentage recoveries were determined to be 98.34, 99.19, 100.18, 102.97 and 99.81%, respectively, at the five concentrations between 5.0 and 100 microm. The limit of quantitation of naproxen was determined to be 0.29 microg/mL, while the detection limit was 64 ng/mL. Four major photoproducts were observed from the HPLC chromatogram using a Panchum PR-2000 reactor which equipped with 8 W x 16 low-pressure quartz mercury lamps as the light source for irradiation of a naproxen sample in methanol. The structures of the photoproducts were confirmed by LC-ESI MS., (Copyright 2005 John Wiley & Sons, Ltd.)

Serum creatinine (SCr) had been considered to be an important predictor of mortality in end-stage renal disease (ESRD) patients at the start of renal replacement therapy (RRT). However, the data were limited about initially extreme azotemia (EA), exclusively defined as blood urea nitrogen (BUN) > or = 300 mg/dL, SCr > or = 30 mg/dL, or both. This retrospective study was conducted to clarify the characteristics and outcome in our EA patients. We had 1682 new ESRD patients from July 1988 to December 1996. With frequency match for age, gender, and starting RRT in the same period, 20 EA patients and 60 controls were included. Fifty percent of our EA patients had unknown etiology. The EA patients had significantly lower prevalence of underlying diabetic nephropathy, and comorbid hypertension. All the EA patients had late referral to nephrologists within 4 weeks before the initiation of RRT, and 90% of them had taken Chinese herbals. The EA group had significantly higher BUN, SCr, and iron storage as well as a higher prevalence of severe anemia, hyperkalemia, hypocalcemia, and acidemia. However, the similar prevalence of cardiomegaly and left ventricular hypertrophy as well as the similar early mortality rate and long-term survival were noted. Age over 40 years, comorbid diabetes mellitus, and hypoalbuminemia were independent predictors of poor survival. Our EA patients had different initial presentations from other uremic ones at the start of RRT. However, the short-term and long-term mortality rates were similar. The lower prevalence of underlying diabetic nephropathy and comorbid hypertension among the EA patients might contribute to their fair outcome.

We described to achieve the local retention of minoxidil which has penetrated the skin with minimization of its absorption into the general circulation and elimination of local irritation induced by propylene glycol. The effect of tocopheryl polyethylene glycol succinate (TPGS) on the penetration flux of minoxidil and its retention in the skin from topical minoxidil formulations consisting of water, alcohol, and polyethylene glycol 400 was characterized by an experimental design of ten solvent formulations in this study. Results show that the addition of TPGS was only able to improve the solubility of minoxidil in those solvent systems containing higher proportions of water and PEG 400, and the extent of improvement was also more profound with the addition of TPGS at concentrations higher than 5%. For those solvent systems containing a higher fraction of alcohol, an insignificant change in minoxidil solubility with increasing added amounts of TPGS was noted even with the tendency to decrease the solubility of minoxidil with higher amounts of TPGS. Increasing the amount of TPGS added gradually increased the flux and the corrected flux from solvent formulations with a lower solubility parameter, but decreased those from solvent systems with a higher solubility parameter. With the addition of TPGS, solvent formulation F6 (alcohol:PEG 400 of 50:50) was demonstrated to be the optimal choice by having an improved local effect and a reduced systemic effect compared to the reference of 2% Regaine((R)). Tocopheryl polyethylene glycol succinate (TPGS) was mainly retained locally in the stratum corneum, and the amount was proportional to the increase in the amount of TPGS added to these ten solvent formulations.

In researches of ketone bodies, D-3-hydroxybutyrate (D-3HB) is usually the major one which has been investigated; in contrast, little attention has been paid to L-3-hydroxybutyrate (L-3HB), because of its presence in trace amounts, its dubious metabolism, and a lack of knowledge about its sources. In the present study we determined the distributions of enantiomers of 3-hydroxybutyrate (3HB) in rat brain, liver, heart, and kidney homogenates, and we found the heart homogenate contained an enriched amount of L-3HB (37.67 microM/mg protein) which generated a significant ratio of 66/34 (D/L). The ratio was altered to be 87/13 in the diabetic rat heart homogenate. We subsequently found this changed ratio of D/L-3HB may contribute to reduce glucose utilization in cardiomyocytes. Glucose utilization by cardiomyocytes with 5 mM of D-3HB was decreased to 61% of the control, but no interference was observed when D-3HB was replaced with L-3HB, suggesting L-3HB is not utilized for the energy fuel as other ketone bodies are. In addition, the reduced glucose utilization caused by D-3HB gradually recovered in a dose-dependent manner with administration of additional L-3HB. The results gave the necessity of taking L-3HB together with D-3HB into account with regard to glucose utilization, and L-3HB may be a helpful substrate for improving inhibited cardiac pyruvate oxidation caused by hyperketonemia.

In this study, topical minoxidil solutions supplemented with TPGS in cosolvent systems of various compositions consisting of water, alcohol, and polyethylene glycol 400 were designed to evaluate the efficacy of promoting hair growth after topical application and the safety in terms of the amount of minoxidil absorbed through the skin into the circulation using C57BL/6J mice as a model. The commercial product of 2% Regaine) was used as the positive control. The role, which sulfotransferase activity plays in hair growth with treatment using minoxidil, was determined as well. The results revealed that the addition of 0.5% TPGS was able to enhance the proliferation of hair, but an increase in the amount of TPGS to 2% led to deterioration in the enhancement of hair growth. At the higher added amount (2.0%) of TPGS, the promotion of hair growth was slightly reduced for both cosolvent formulations F1 (100% water) and F3 (100% PEG 400), whereas it was reduced to a greater extent for the cosolvent formulations F8-F10. In comparison, the influences of cosolvent compositions with TPGS amounts of 0.0 and 2.0% on the promotion of hair growth were similar. On the contrary, variability in the promotion of hair growth by different solvent formulations was minimal when the added amount of TPGS was 0.5%. In general, a relationship between hair growth and sulfotransferase activities after topical application of 2% Regaine and minoxidil formulations containing various amounts of TPGS was not demonstrated. Plasma concentrations of minoxidil with 2% Regaine were found to be greater than those of 2% minoxidil in those cosolvent formulations containing various amounts of TPGS, while showing insignificant differences among those 10 cosolvent formulations with a fixed amount of TPGS. A tendency for the plasma concentration of minoxidil to increase after the topical administration of minoxidil formulations containing the higher amount of TPGS (2%) was noted.

Indomethacin is used as an anti-inflammatory drug and a nonselective cyclooxygenase inhibitor. When indomethacin in methanol was photo-irradiated with an Hg lamp, methyl ester, ethyl ester, and gamma-lactone derivatives of indomethacin were produced. In the present study, we found that the methyl ester derivative of indomethacin (M-IN) could more potently inhibit prostaglandin E(2) (PGE(2)) and nitric oxide (NO) production and inducible nitric oxide synthase (iNOS) and cyclooxygenase 2 (COX 2) protein expression from lipopolysaccharide (LPS)-stimulated RAW 264.7 cells than indomethacin, similar to the effect of a non-steroidal anti-inflammatory drugs (NSAID). On the other hand, the results showed that M-IN with an IC(50) value maintained at 36.9 microg/ml for 12 h exhibited stronger cytotoxicity than ethyl ester, gamma-lactone derivatives of indomethacin, and indomethacin in promyelocytic leukemia HL-60 cells. Moreover, a series of biochemical analyses determined that M-IN caused apoptotic bodies, DNA fragmentation, and enhanced PARP and pro-caspase 3 degradation in HL-60 cells. These above results indicate that the photosynthesized product, M-IN, had stronger anti-inflammatory effects in LPS-stimulated RAW 264.7 cells and cytotoxicity effects in HL-60 cells than the parent drug, indomethacin.

Fusarium head blight (FHB) or scab caused by Fusarium species is an economically important disease on small grain cereal crops worldwide. Accurate assessments of the pathogenicity of fungal isolates is a key obstacle toward a better understanding of the Fusarium-wheat scab system. In this study, a new laboratory method for inoculation of wheat coleoptiles was developed, which consists of cutting off the coleoptile apex, covering the cut apex with a piece of filter paper soaked in conidial suspension, and measuring the lengths of brown lesions 7 days post inoculation. After coleoptile inoculation, distinct brown lesions in the diseased stems were observed, in which the presence of the fungus was verified by PCR amplification with F. graminearum Schwable-specific primers. Coleoptile inoculation of six wheat varieties indicated that a highly susceptible wheat variety was more suitable as a differentiating host for the pathogenicity assay. Analysis of the coleoptiles inoculated with a set of 58 different isolates of F. graminearum showed a significant difference in the lengths of the lesions, forming the basis by which pathogenicity of the isolates was assessed. Field inoculation of florets of three wheat varieties over 2 years revealed significant differences in pathogenicity among the 58 isolates, and that the highly resistant and highly susceptible wheat varieties were more appropriate and stable for pathogenicity assessment in field trials. Comparative analyses of eight inoculation experiments of wheat with 58 F. graminearum isolates showed significant direct linear correlations (P<0.001) between coleoptile and floret inoculations. These results indicate that the wheat coleoptile inoculation is a simple, rapid and reliable method for pathogenicity studies of F. graminearum in wheat.

Based on the intergenic sequences of Tri5-Tri6 genes involved in the mycotoxin pathways of Fusarium species, a generic PCR assay was developed to detect a 300 bp fragment of deoxynivalenol (DON)-chemotypes and a 360 bp sequence of nivalenol (NIV)- chemotypes of Fusarium graminearum. Mycotoxin chemotypes identified by the PCR assays were confirmed by the chemical analyses of HPLC or GC/MS. Further analysis of 364 F. graminearum isolates from 12 provinces of China showed that 310 were DON-chemotypes and 54 were NIV-chemotypes. Sequence analyses revealed that DON-chemotypes display more variations than NIV-chemotypes. This PCR assay could be used to detect mycotoxin-producing Fusarium-species and may thus help to develop strategies to avoid or reduce mycotoxin contamination of cereals. Also this assay may provide useful alternatives to antibody-based mycotoxin tests.

A sample of 10 mm zomepirac in methanol was photo-irradiated with a Hanovia 200 W high-pressure quartz Hg lamp for 14 days. In total, four photoproducts were observed from the HPLC chromatogram. The preparative HPLC included an YMC-Pack Pro C18 column (250 x 20 mm i.d.), a mobile phase of CH3CN-CH3OH-1%HOAc (10:60:30, v/v/v), and UV detection at 254 nm. The most probable structures of the four photoproducts were determined by LC-MS. Two major photoproducts were separated, and their structures were further confirmed by the spectroscopic methods. A reaction scheme of zomepirac was proposed that the photochemical reaction routes occur mainly via bond fission between carbonyl-pyrrolyl groups (alpha-cleavage of a ketone), and decarboxylation followed by oxidation with singlet oxygen to produce an aldehyde., (Copyright 2004 John Wiley & Sons, Ltd.)

A high-performance liquid chromatography (HPLC) method using silica column eluted with aqueous solvent mobile phase containing triethylamine (TEA) and acetic acid (ACH) at trace percentages was characterized for the analysis of basic compounds. The key mechanism of this system is ion-exchange accompanying interaction of silanol groups. The increase in the ACH concentration in the mobile phase minimizes the ionization of the silanol group, leading to reduced retention time. However, the greater extent of ionization of silanol caused by the increase of TEA concentration helps to retain basic compounds in the column. Further, the protonated TEA that is positively charged also competes for the ionized silanol group with basic compounds, resulting in the modification of retention time. On the other hand, the retention becomes longer with increasing proportion of either organic or aqueous solvent in mobile phase, and partial replacement of methanol with acetonitrile.

A high-performance liquid chromatographic (HPLC) assay method for assessing the degradation of tolmetin (TLM) is developed and validated under acidic, basic, and photoirradiated conditions. The HPLC method includes an Inertsil 5 ODS-3V column (250- x 4.6-mm i.d.), guard column of Inertsil 7 ODS-3V (50- x 4.6-mm i.d.), mobile phase of CH(3)OH-1% HOAc (64:36, v/v), and UV detection at 254 nm. The developed method satisfies the system suitability criteria, peak integrity, and resolution for the parent drug and its degradants. The established assay method exhibits good selectivity and specificity suitable for stability measurements. From the intra- and interday tests of six replicates, the coefficients of variation are between 0.20% and 1.77% for the former, and 0.12% and 3.40% for the latter. Recoveries are found to be 98.7-101.7%. TLM is determined to be more reactive when exposed to light and acidic conditions, yet TLM is stable in a basic medium. A kinetic study of the photodegradation of TLM shows that it follows an apparent first-order reaction in three alcoholic solvents.

An improved HPLC method was developed for the determination of sildenafil concentrations in plasma. Analysis of sildenafil in plasma samples was simplified by utilizing a one-step liquid-liquid extraction after alkaline treatment of only 1 ml of plasma. The lower limit of quantitation was 10 ng/ml with a coefficient of variation of less than 20%. A linear range was found to exist from 10 to 1000 ng/ml. This HPLC method was validated with precisions (coefficient of variation, C.V.) for inter- and intra-day runs of 0.41-11.15% and 0.36-8.05%, respectively, and accuracies (the relative error of the mean, REM) for inter- and intra-day runs of -8.72-6.81% and 0.41-11.15%, respectively. A bioavailability study of sildenafil was performed on one normal healthy male volunteer by analyzing sildenafil plasma concentrations with this validated HPLC method. Results demonstrated that this HPLC method is appropriate for applications to bioavailability studies of sildenafil. In addition, an example of the influence of the co-administration of grapefruit juice on sildenafil pharmacokinetics in a healthy volunteer is presented.

A solution of carprofen in methanol at a concentration of 2.74 x 10(-2) mg/mL is subjected to photoirradiation using a Hanovia 200-W high-pressure Hg lamp for 9 h. In total, seven photodegradation products are separated, and their quasimolecular ions are subsequently determined online using a liquid chromatography (LC)-electrospray ionization (ESI)-mass spectrometry (MS) method. The high-performance LC consists of an Inertsil 5 ODS-80A (2.1- x 150-mm) column. The mobile phase is initially CH(3)CN. NH(4)OAc (20mM in de-ionized H(2)O) is 43:57 (v/v), and after 14 min it is CH(3)CN. NH(4)OAc (20mM in de-ionized H2O) is 54: 46 (v/v). The UV detector was set at 260 nm. The parameters of LC-MS for mass determination involves an atmospheric pressure ionization electron spray interface with a negative mode of polarity (ESI(-)). The chemical structures of the degradants are elucidated based on the mass-to-charge ratio of the quasimolecular ions and the molecular weight changes by comparison with the parent drug (carprofen). The degradation proceeds via an initial dechlorination. A dechlorination or esterification reaction is competed with decarboxylation. This finding is in accordance with our previously reported result of first order photodecomposition kinetics for carprofen.

A rapid, sensitive, and accurate stability-indicating high-performance liquid chromatographic assay method for determining the degradation of carprofen (CPF) is developed and validated under acidic, basic, or photo-irradiated conditions. The analysis is monitored with a Cosmosil 5C18-AR column using a mobile phase of CH3CN-H2O-AcOH (50:49:1, v/v/v) at 260 nm. The developed method satisfies the system suitability criteria, peak integrity, and resolution among the parent drug and its degradation products. The results indicate that the established assay method shows good selectivity and specificity suitable for stability measurements of CPF. CPF is found to be more sensitive to exposure to light and in acidic conditions, but it is stable in a basic medium. The kinetic study of the photodegradation of CPF follows an apparent first-order reaction in a variety of solvents. The solvent effects on the rates of degradation are in the decreasing order of chloroform > dichloromethane > methanol > ethanol > 2-propanol, which is irrelevant to the dielectric constant epsilon. However, the hydrogen-donating ability of the solvents is essential to the photochemical decomposition of CPF. A plot of log k versus the Kirkwood function exhibits a linear relationship in aqueous ethanolic solutions, which implies that degradation proceeds via an ionic mechanism.