Comparative ecotoxicity of single and binary mixtures exposures of nickel and zinc on growth and biomarkers of Lemna gibba

The aim of this study was to compare the ecotoxicity of nickel (Ni) and zinc (Zn) assayed as single and as binary mixture. In addition, how were affected the population growth rates and oxidative stress biomarkers, comparing single to binary exposures. The toxicity tests were performed on Lemna gibba using a 7-day test. All calculations were made using measured total dissolved metal concentrations. IC50-7d, based on growth rate calculated on frond number and fresh weight, were 2.47/3.89 mg/L, and 76.73/76.93 mg/L, for Ni and Zn, respectively. Single metals affected plant growth following a non-linear concentration–response relationship. LOEC values for each metal were obtained at 0.92 and 20.1 mg/L for Ni and Zn, respectively. Biomarkers of the antioxidant response like Catalase (CAT; EC 1.11.1.6), ascorbate peroxidase (APOX; EC 1.11.1.11) and guaiacol peroxidase (GPOX; EC 1.11.1.7) activities in single metals assays were higher than controls, but when similar concentrations were added as mixtures, that increase was reduced and inhibition with respect to the control was observed for GPOX. APOX showed the highest activity. The concentration addition (CA) approach was evaluated and resulted in a correct predictor of Ni-Zn mixture toxicity on Lemna gibba. This was made comparing the EC50 and LOEC, measured taking the growth rate as endpoint, with those expected values according to the CA model. However, the measured biomarkers indicating a positive response to free radicals did not fit to concentration addition model when assayed in the binary mixture. Also, the main activity response of these was observed within a range of concentrations below the LOEC values for the mixture.

     

Antimicrobial activity of isolate HL-12 against Clavibacter michiganensis subsp. michiganensis in the presence of cadmium

An actinomycete, strain HL-12, that was isolated from a farmland on the Huajiachi campus of Zhejiang University was capable of inhibiting the growth of Clavibacter michiganensis subsp. michiganensis (Cmm) and was identified as a member of Streptomyces. Its antimicrobial activity against Cmm was measured using the agar plate sensitivity method in pure culture and evaluated by the inhibition ratio of Cmm in soil. The inhibitory activity of strain HL-12 against Cmm following exposure to low concentrations of Cd was greater than the inhibitory activity following exposure to high concentrations of Cd both in liquid culture and in soil. A stronger inhibition was also seen following a 24 h preculture in the presence of Cd in liquid culture. The growth of Cmm in soil was stimulated at low concentrations of Cd (<5.0 mg Cd kg⁻¹ dry soil) but inhibited when cultured in high concentrations of Cd (5.0 and 10.0 mg Cd kg⁻¹ dry soil). A higher inhibition ratio of strain HL-12 against Cmm, which was over 40% after soil incubation for 2 weeks, was observed following exposure to low concentrations of Cd (<5.0 mg Cd kg⁻¹ dry soil).     

The physiological response and sub-cellular localization of lead and cadmium in Iris pseudacorus L.

The seedling development and physiological responses of Iris pseudacorus L. to Pb and Cd and their combination were studied for 28 days liquid culture and sub-cellular localization of Pb and Cd in the root tip cells treated with 2,070 mg L⁻¹ Pb and 1,000 mg L⁻¹Cd for 16 days sand culture was evaluated. Results showed that the dry weights (DWs) of shoots and roots of I. pseudacorus were significantly decreased at 500 mg L⁻¹Pb and 25 mg L⁻¹Cd + 500 mg L⁻¹Pb treatments and the root DWs under all treatments were significantly decreased in comparison with that of control. The concentrations of Chla in the leaves were decreased at all treatments, while, the concentrations of Chlb and total carotenoids were not significantly decreased under 25 mg L⁻¹Cd and 25 mg L⁻¹Cd + 500 mg L⁻¹Pb treatments. The MDA and proline concentrations and POD activities in the shoots and roots were increased under treatments of 500 mg L⁻¹Pb and 25 mg L⁻¹Cd + 500 mg L⁻¹Pb, but POD activities in the shoots and roots and MDA concentrations in the shoots were significantly decreased at 25 mg L⁻¹ Cd treatment. The results of sub-cellular localization of Pb and Cd showed that numerous Pb deposits were found on the inner surface of died cell walls in the cortex treated with 2,070 mg L⁻¹ Pb and Cd deposits were found in the cell wall treated with 1,000 mg L⁻¹ Cd. Pb and Cd deposits were not found in the cytoplasm. The results indicated that POD and proline showed strong beneficial properties against Pb and Cd stress and there were some mechanisms keeping most cells with normal activities in the plant from Pb toxicity by sacrificing a few cells that accumulated a large amount Pb. Sub-cellular localizations of Pb and Cd in the root tip cells of I. pseudacorus were little difference with the localizations in other species of Iris in the previous studies.     

Influence of combined treatment with zinc and selenium on cadmium induced testicular pathophysiology in rat

The present study was conducted to evaluate the potential benefit of combined treatment with zinc (Zn) and selenium (Se) in reversing cadmium (Cd)-induced testicular pathophysiology compared to Se or Zn treatment alone in rats. For this purpose, male rats received either tap water, Cd, Cd + Zn, Cd + Se or Cd + Zn + Se in their drinking water, for 35 days. Cd exposure caused a significant decrease in plasma and testicular concentrations of Se and Zn which was accompanied by decreased plasma testosterone level, sperm count and motility, enzymatic activity of superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GSH-Px) as well as by increased lipid peroxidation (as malondialdehyde, MDA). With Se or Zn administration, during exposure to Cd, only partial corrective effects on depletion of testicular and plasma Se and Zn levels, sperm characteristics and oxidative stress have been observed. The combined treatment of Cd-exposed animals with Se and Zn assured a more significant decrease in plasma and testicular Cd concentrations and a more efficient protection against the observed testicular damage as evidenced by the total prevention of both Se and Zn deprivation and by the entire restoration of the sperm motility and the testicular antioxidant status.     

Genotoxicity response of Vicia faba seedlings to cadmium in soils as characterized by direct soil exposure and micronucleus test

To overcome the drawbacks of the Vicia faba root tip micronucleus test in soil using the solution extract method, we conducted a potting experiment by direct soil exposure. Cadmium was spiked into 3 typical soils (brown soil, red soil, and black soil) to simulate environmental concentrations (0.625, 1.25, 2.5, 5, and 10 mg kg−1). Multiple Vicia faba tissues (primary root tips, secondary root tips, and leaf tips) were sampled, and mitotic index (MI), chromosome aberration frequency (CA), and micronucleus frequency (MN) were used as endpoints after a seedling period of 5 days. The results showed a response between Cd concentrations and multiple sampling tissues of Vicia faba, and the secondary root tips responded to Cd stress the most, followed by primary root tips and leaf tips. Soil physicochemical properties (e.g., pH, total phosphorus, total organic carbon, etc.) influenced the genotoxicity of Cd, and pH was the dominant factor, which resulted in the genetic toxicity response of Cd in soils in the order: red soil > brown soil > black soil. The lowest observable effect concentration (LOEC) of Cd was 1.25 mg kg−1 for both brown soil and red soil and 2.5 mg kg−1 for black soil. In view of this, we suggested that soil properties should be considered in evaluating genotoxicity risk of Cd in soil, especially with soil pH range, and the secondary root tips should be taken as suitable test tissues in the MN test due to its more sensible response feature to Cd stress in soil.     

Morphine-induced Neuroimmunomodulation in Murine Visceral Leishmaniasis: The Role(s) of Cytokines and Nitric Oxide

Opioid modulation of host resistance to infectious diseases is well documented; however, not much is known during visceral leishmaniasis (VL). Low doses of morphine, administered subcutaneously in Leishmania donovani-infected BALB/c mice, on days 0 and +15, significantly (p < 0.05) suppressed (1 mg/kg/day) or even sterile-cleared (2 mg/kg/day) the infection; paradoxically, high doses (10 and 30 mg/kg/day) exacerbated the infection. In vitro, low concentration (1 × 10⁻⁹ and 1 × 10⁻¹¹ M) morphine treatment of L. donovani-infected mouse peritoneal macrophages (PM), endowed them with significant (p < 0.05) leishmanicidal activity, whereas a high-concentration (1 × 10⁻⁵ M) treatment augmented intramacrophage parasite growth. Naloxone pre-treatment of infected-mice (4 mg/kg × 2) and of infected-PM (1 × 10⁻⁵ M), blocked only the morphine low dose/concentration-induced protective effect. The splenocytes from protected mice and morphine low concentration-treated infected-PM, elaborated significantly (p < 0.05) enhanced levels of interleukin-12, interferon-γ, tumor necrosis factor-α, granulocyte-macrophage colony-stimulating factor and nitrite in the culture medium; a high dose/concentration suppressed their elaboration. Curiously, only morphine high dose/concentration-treated infected mice splenocytes and infected PM, produced significantly (p < 0.05) increased quantity of transforming growth factor-β1. Aminoguanidine, significantly (p < 0.05) blocked the morphine low dose/concentration-induced protective effect, in vivo and in vitro. This first study demonstrates dose-dependent biphasic modulatory effects of morphine in L. donovani-infected mice and PM, in vitro, apparently via nitric oxide-dependent mechanisms. These results thus demonstrate the implications of opiate abuse on the efficacy assessment of antileishmanial drugs and vaccines, and on the reactivation of latent VL in areas where both drug abuse and VL are rampant. Disclaimers: nil Sources: Senior Research Fellowship to P. S. National Eligibility Test, Senior Research Fellow, The Council of Scientific and Industrial Research, New Delhi, India. Drugs. morphine sulphate (Government Opium and Alkaloid Factory, Ghazipur, India). Meeting presentation. part of the work presented in 12th Society on NeuroImmune Pharmacology Conference, Santa Fe, New Mexico, USA, April 05–9, 2006.     

An iron-deficient diet stimulates the onset of the hepatitis due to hepatic copper deposition in the Long-Evans Cinnamon (LEC) rat

To study effects of dietary Cu and Fe levels on the onset of hepatitis in Long-Evans Cinnamon (LEC) rats, female rats (40 days old) were fed a semipurified diet containing 0.1 or 10 mg Cu/kg and 1.5 or 150 mg Fe/kg in a 2 × 2 factorial arrangement for 35 days. At 75 days after birth, LEC rats (+Cu−Fe) fed a Cu-sufficient but Fe-deficient diet (Cu, 10 mg/kg; Fe, 1.5 mg/kg) showed jaundice, with lethargy, anorexia, and malaise. The biochemical variables relating to liver function were significantly increased compared to three other groups, a Cu- and Fe-deficient (−Cu−Fe) group, a Cu-deficient but Fe-sufficient (−Cu+Fe) group, and a Cu and Fe sufficient (+Cu+Fe) group. Furthermore, the +Cu−Fe rat liver showed massive necrosis with huge nuclei. The other three groups presented no biochemical and histological findings of hepatitis. Hepatic Cu and metallothionein concentrations were 289 ± 87 (mean ± SD) μg/g liver and 8.7 ± 1.8 mg/g liver, respectively, in the +Cu−Fe rats. However, in the +Cu+Fe group the values were 196 ± 28 μg Cu/g liver and 10.8 ± 1.0 mg/g liver. Hepatic Fe deposition was not influenced significantly by the dietary Cu level. The +Cu−Fe group with jaundice showed the highest free Cu concentration in the liver among the four groups, but the hepatic free Fe concentration was similar to those in the −Cu+Fe and +Cu+Fe groups. Our results indicate that an Fe-deficient diet enhances the deposition of hepatic Cu due to increased absorption of Cu from the gastrointestinal tract. This deposition stimulated the onset of hepatitis. Received: 11 March 1999 / Accepted: 11 June 1999     

Pharmacokinetics and safety of candesartan cilexetil in subjects with normal and impaired liver function

Objective: The influence of liver disease on the pharmacokinetics of candesartan, a long-acting selective AT₁ subtype angiotensin II receptor antagonist was studied. Methods: Twelve healthy subjects and 12 patients with mild to moderate liver impairment received a single oral dose of 12 mg of candesartan cilexetil on day 1 and once-daily doses of 12 mg on days 3–7. The drug was taken before breakfast. Serial blood samples were collected for 48 h after the first and last administration on days 1 and 7. Serum was analyzed for unchanged candesartan by HPLC with UV detection. Results: The pharmacokinetic parameters on days 1 and 7 revealed no statistically significant influence of liver impairment on the pharmacokinetics of candesartan. Following single dose administration on day 1, the␣mean␣C_max was 95.2 ng · ml⁻¹ in healthy subjects and 109 ng · ml⁻¹ in the patients. The AUC_0−∞ was␣909 ng.h · ml⁻¹ in healthy volunteers and 1107 ng.h · ml⁻¹ in patients and the elimination half-life was 9.3 h in healthy volunteers and 12 h in the patients. At steady state on day 7, mean C_max values were similar in both groups (112 vs 116 ng · ml⁻¹); the AUC_τ was 880 ng.h · ml⁻¹ in healthy subjects and 1080 ng.h · ml⁻¹ in patients while the elimination half-life was 10 h in healthy subjects and 12 h in the patients with liver impairment. The AUC_0−∞ on day 1 was almost identical to the AUC_τ on day 7. A moderate drug accumulation of 20%, which does not require a dose adjustment, was observed following once-daily dosing in both groups. No serious or severe adverse events were reported. Conclusion: Mild to moderate liver impairment has no clinically relevant effect on candesartan pharmacokinetics, and no dose adjustment is required for such patients. Received: 24 November 1997 / Accepted in revised form: 18 February 1998     

Combined effects of okadaic acid and cadmium on lipid peroxidation and DNA bases modifications (m5dC and 8-(OH)-dG) in Caco-2 cells

Okadaic acid (OA) is a marine toxin, a tumour promoter and an inducer of apoptosis. It mainly inhibits protein-phosphatases, protein synthesis and enhances lipid peroxidation. Cadmium (Cd) is known to be carcinogenic in animals and humans (group 1 according to the International Agency for Research on Cancer (IARC) classification). Cd also induces oxidative stress in living organisms. Since they are sometimes found simultaneously in mussels, we have evaluated in the present investigation, the lipid peroxidation, as malondialdehyde (MDA) production, in the variation of the ratios of 8-(OH)-dG/10⁵dG and m⁵dC/ (dC + m⁵dC) induced by OA and/or Cd in Caco-2 cells. When cells were treated exclusively by OA (15 ng/ml) or Cd (0.625 and 5μg/ml) for 24 h, protein synthesis was inhibited (by 42 ± 5%, 18 ± 13%, and 90 ± 4% respectively) while MDA production was 2235 ± 129, 1710 ± 20, and 11496 ± 1624 pmol/mg protein respectively. In addition, each toxicant induced modified bases in DNA; increases in oxidised bases and methylated dC. The combination of OA and cadmium was more cytotoxic and caused more DNA base modifications; the ratio m⁵dC/(m⁵dC+dC) was increased from 3 ± 0.15 to 9 ± 0.15 and the ratio 8-(OH)-dG/10⁵ dG also (from 36 ± 2 to 76 ± 6). The combination of OA and Cd also increased the level of MDA (16874 ± 2189 pmole/mg protein). The present results strongly suggest that DNA damage resulting from the oxidative stress induced by these two toxicants may significantly contribute to increasing their carcinogenicity via epigenetic processes. Received: 28 September 1999 / Accepted: 10 January 2000     

Effect of verapamil on tachycardia-induced early cellular electrical remodeling in rabbit atrium

We investigated the effects of a 7-day verapamil pretreatment (VPT, 7.5 mg/kg bodyweight subcutaneously every 12 h) on ionic currents and molecular mechanisms underlying tachycardia-induced early electrical remodeling after 24-h rapid atrial pacing (RAP, 600 bpm) in rabbit atrium. Animals were divided into four groups (n = 6 each group): control (not paced, no verapamil), paced only, verapamil only and verapamil and paced, respectively. VPT doubled ICa,L [7.0 ± 0.7 pA/pF (control) vs 14.2 ± 0.6 pA/pF (verapamil only)]. RAP reduced ICa,L by 48% to 3.6 ± 0.7 pA/pF (paced only). RAP did not affect ICa,L in verapamil-treated animals and averaged 15.3 ± 0.2 pA/pF (paced and verapamil). RAP resulted in a significant decrease of the expression of the α1c subunit (−24.7%) and the β2A subunit (−13.3%), respectively. VPT led to a similar alteration of subunit expression as RAP [“control” vs “verapamil only”, decrease of α1c subunit (−25.4%), but no significant change in β2A subunit expression]. However, after VPT, further diminishment of α1c and β2A subunit expression after rapid atrial pacing was absent. (“verapamil” vs “verapamil and paced”, n = 6 both groups). RAP decreased Ito [−45%, 51.5 ± 3.9 pA/pF (control) vs 26.8 ± 1.5 pA/pF (paced only)] and was not influenceable by VPT. IK1 was neither affected by RAP nor verapamil pretreatment. Downregulation of α1c and β2A subunit expression and the resulting decay of ICa,L current densities were prevented by verapamil. However, these effects are abolished by multiple other adverse effects of verapamil on atrial electrophysiology.     

Lack of a pharmacokinetic interaction between atovaquone and proguanil

Objective: To assess the magnitude of the putative effect of atovaquone on the pharmacokinetics of proguanil and to determine whether the pharmacokinetics of atovaquone are affected by concomitant administration of proguanil, with both drugs administered for 3 days to healthy adult volunteers. Methods: This was an open-label, randomized, three-way cross-over study, in which 18 healthy volunteers received 400 mg proguanil, 1000 mg atovaquone and 1000 mg atovaquone + 400 mg proguanil. Each treatment was given once daily for 3 days with a 3-week wash-out period between each occasion. For the assay of proguanil, cycloguanil and atovaquone, blood was sampled before dosing and at regular intervals over 8 days when proguanil was given, and over 17 days when atovaquone was given. Results: The geometric mean of the area under the atovaquone plasma concentration-time curve calculated from 0 to 24 h after the last dose (AUC_0→24h) was 180 μg · ml⁻¹ · h following administration of atovaquone alone and 193 μg · ml⁻¹ · h following atovaquone with proguanil. The geometric mean AUC_0→24h for proguanil was 6296 ng · ml⁻¹ · h after proguanil alone and 5819 ng · ml⁻¹ · h following proguanil with atovaquone. The corresponding values for the metabolite cycloguanil were 1297 ng · ml⁻¹ · h and 1187 ng · ml⁻¹ · h, respectively. The geometric mean elimination half-life (t_1/2) of atovaquone was 57.1 h when given alone and 59.0 h when administered together with proguanil. The corresponding geometric mean values of t_1/2 for proguanil were 13.7 h and 14.5 h. Exploratory statistical analysis showed no important gender effects on the pharmacokinetics of atovaquone, proguanil, or cycloguanil. Conclusion: The pharmacokinetics of atovaquone and proguanil and its metabolite, cycloguanil, were not different when atovaquone and proguanil were given alone or in combination. Received: 14 October 1998 / Accepted in revised form: 8 February 1999     

Measurement of furancarboxylic acid, a candidate for uremic toxin, in human serum, hair, and sweat, and analysis of pharmacological actions in vitro

3-Carboxy-4-methyl-5-propyl-2-furanpropionic acid (CMPF), a candidate for uremic toxin, was measured in human hair for examining a possible utility as indicator of renal dysfunction. The serum concentration of CMPF was much higher (32.3 ± 2.7 μg/ml, n=17; mean ± SEM) in uremic patients aged 40–55 years receiving hemodialysis treatment than in healthy younger subjects (3.61 ± 0.19 μg/ml, n=22), aged 18–23 years. However, the hair concentration of CMPF tended to be lower in the patients (6.8 ± 1.7 ng/10 mg hair) than in the healthy younger subjects (15.8 ± 4.5 ng/10 mg) and was significantly lower than that in the healthy age-matched subjects (22.4 ± 5.3 ng/10 mg, n=12), aged 40–47 years. Since CMPF was measurable in the sweat (4.4 ± 3.7 ng/mg) collected from six out of seven healthy subjects examined, it was suggested that the contribution of sweat to the measurement of CMPF in hair was considerable. The fact that the uremic patients undergoing hemodialysis therapy had less sweat than healthy subjects may explain the lower concentration of CMPF in the patients' hair. The pathophysiological roles of CMPF in the body were attempted to be explored by using excised guinea pig organs, and human platelets and neutrophils. CMPF showed no remarkable effects in the concentration range of ≤10⁻⁴ M except for only slight suppression of spontaneous contracture of guinea pig tenia coli at 10⁻⁴ M. As far as the organs and tissues examined in the present study are concerned, the biological activity of CMPF itself, if any, may be very weak. Precaution should be taken against the delivery of a substance through sweat to hair when a small amount of substance is attempted to be measured in hair by employing a sensitive analytical method. Received: 14 September 1999 / Accepted: 3 November 1999     

Evaluation of involvement of testicular metallothionein gene expression in the protective effect of zinc against cadmium-induced testicular pathophysiology in rat

To investigate the effects of exposure to Cd and Zn on testicular MT-1 and MT-2 gene expression and evaluate their involvement in Zn protection against Cd-induced testicular pathophysiology, male rats received either tap water, Cd or Cd + Zn in their drinking water for 35 days. Cd induced histopathological changes in testicular tissues were accompanied by decreased plasma testosterone level, plasma and testicular Zn concentrations, oxidative stress, and by increased MT-1 and MT-2 gene expression. Co-treatment with Cd and Zn reversed the Cd-induced decrease testosterone level and SOD activity, decreased testicular Cd accumulation and partially restored Cd-induced histological changes, lipid peroxidation, and Zn depletion. The increase of testicular MT-1 and MT-2 gene expression under Cd influence was significantly reduced in Cd + Zn group. These data suggest that Zn enhances the protection against Cd-induced testicular pathophysiology through non-MT gene expression mechanisms but essentially by preventing Cd accumulation, Zn deprivation and by ameliorating the testicular antioxidant status.     

Total and free mycophenolic acid and its 7-O-glucuronide metabolite in Chinese adult renal transplant patients: pharmacokinetics and application of limited sampling strategies

Objective This study aimed to investigate the pharmacokinetic characteristics of total and free mycophnolic acid (MPA) and its 7-O-glucuronide metabolite (MPAG) in Chinese renal transplant recipients. In addition, limited sampling strategies were developed to estimate the individual area under concentration curve (AUC) of total and free MPA. Methods Total and free MPA and MPAG concentrations were determined by high performance liquid chromatography. Whole 12-h pharmacokinetic profiles were obtained on the 10th day after operation in 12 adult Chinese de novo renal transplant recipients administrated with mycophenolate mofetil (MMF, 750 mg bid), cyclosporine and corticosteroids. Limited sampling strategies with jackknife technique, a resampling method, and Bland-Altman analysis were employed to develop equations to estimate total and free MPA AUC. Results The pattern of total and free MPA and MPAG plasma concentration-time curves in the cohort of patients taking lower doses of MMF was consistent with previous reports of Caucasian patients taking MMF 1 g bid, except that dose-normalized exposure of total and free MPAG was much lower in the current study than in those of the Caucasians. The mean C _max and AUC_0–12h of total and free MPA were 9.4 ± 3.4 mg/L, 20.2 ± 6.5 mg·h/L and 0.4 ± 0.4 mg/L, 0.7 ± 0.5 mg·h/L, respectively, whereas mean C _max and AUC_0–12h of total and free MPAG were 97.3 ± 32.6 mg/L, 656.0 ± 148.0 mg·h/L and 29.9 ± 8.5 mg/L, 222.0 ± 58.1 mg·h/L respectively. The mean fractions of free MPA and MPAG were 3.5 ± 2.0 and 34.6 ± 8.0%, respectively. No determinant was identified to influence the pharmacokinetics of total and free MPA and MPAG or the free fraction of MPA and MPAG. The combinations of C _2h−C _4h and C _1h-C _2h-C _3h were the best to estimate free and total MPA AUC_0–12h respectively, whereas the combination of C _2h-C _3h-C _4h and C _1h-C _2h-C _4h was the best to estimate both simultaneously. Conclusion This is the first time that the pharmacokinetics profile of total and free MPA and its main metabolite MPAG has been examined in Chinese adult renal transplant patients. The limited sampling strategies proposed to estimate individual free and total MPA AUC could be useful in optimizing patient care.     

Unexpected effect of concomitantly administered curcumin on the pharmacokinetics of talinolol in healthy Chinese volunteers

Objective To investigate the effect of concomitantly administered curcumin on the pharmacokinetics of the β1 adrenoceptor blocker talinolol. Methods The study was conducted in a self-controlled, two-period experiment with a randomized, open-labeled design, using 12 healthy volunteers and a wash out period of 1 week between the administration of a single oral dose of 50 mg talinolol and the concomitant administration of curcumin (300 mg day⁻¹ for 6 days) and a single oral dose of 50 mg talinolol on the seventh day. Concentrations of talinolol were measured in plasma by high-performance liquid chromatography-electrospray ionization mass spectrometry. Non-compartmental analysis was used to characterize talinolol plasma concentration-time profiles, all pharmacokinetic parameters were calculated using DAS (ver. 2.0) software, and comparisons of mean values were analyzed by the Wilcoxon signed rank test. Differences were considered to be significant at p < 0.05 (two-sided test). Results The consumption of curcumin for 6 days reduced the area under the curve (AUC) from predose to infinity () of talinolol from 1860.0 ± 377.9 to 1246.0 ± 328.2 ng.h mL⁻¹, the highest observed concentration values (C_max) were significantly decreased from 147.8 ± 63.8 to 106.4 ± 39.9 ng mL⁻¹, and the CL/F was increased from 27.9 ± 5.5 to 43.1 ± 13.4 L.h⁻¹ (p < 0.05). There was no significant difference in sampling time for C_max (t_max) and elimination half-life (t_1/2) values between the two periods (p > 0.05). The interindividual variability in AUC_0–60 and C_max of talinolol was comparable in two study periods; the coefficient of variance (CV) of AUC_0–60 and C_max was 26 and 40% after curcumin versus 21 and 43% after talinolol alone, respectively. Conclusion We suggest that the reduced bioavailability of talinolol is most probably due to the low intraluminal curcumin concentration, or possibly due to the upregulation of further ATP-binding cassette transporters, such as MRP2, in different tissues.     

CYP2C19 genotypes and omeprazole metabolism after single and repeated dosing when combined with clarithromycin

Objectives: Omeprazole is metabolized mainly by CYP2C19 which has two major mutations (CYP2C19*2 in exon5 and CYP2C19*3 in exon4) associated with the poor metabolizer (PM) phenotype. The aim of this study was to examine the relationship between genetic polymorphism of CYP2C19 and metabolism of omeprazole administrated as a single dose or as repeated-doses, which were in both cases co-administered with clarithromycin. Methods: Twelve healthy Japanese subjects were typed for CYP2C19 polymorphism. In the single-dose study, plasma levels of omeprazole and its metabolites were measured for 24 h after administration of 20 mg omeprazole and 400 mg clarithromycin to six healthy Japanese subjects. In the repeated-dose study, plasma levels of omeprazole and its metabolites were measured after repeated oral administration of 20 mg omeprazole and 400 mg clarithromycin twice daily for 6 days and then after 20 mg omeprazole and 400 mg clarithromycin once on the 7th day to the other 6 healthy Japanese subjects. Results: In the single-dose study, the areas under the plasma concentration-versus-time curve (AUCs) of omeprazole of homozygotes for the wild-type allele (*1/*1 n = 2), heterozygotes (n = 3) for the CYP2C19*2 (*1/*2) or for the CYP2C19*3 (*1/*3) and heterozygote (n = 1) for the two defects (*2/*3) were on average 450, 1007 and 6710 ng · h⁻¹ · ml⁻¹, respectively. The ratios of AUCs of omeprazole/5-hydroxyomeprazole for *1/*1, *1/*2 or *1/*3 and *2/*3 were 1, 2 and 30, respectively. In the repeated-dose study, the AUCs of omeprazole for *1/*1, *1/*2 or *1/*3 and *2/*3 were 4041 (n = 2), 3149 (n = 3) and 6684 (n = 1) ng · h⁻¹ · ml⁻¹, respectively. The ratios of AUCs of omeprazole/5-hydroxyomeprazole for *1/*1, *1/*2 or *1/*3 and *2/*3 were 7, 11 and 30, respectively. In the repeated-dose study, the AUC of omeprazole of *1/*1 genotypes was nine-fold higher, that of *1/*2 and *1/*3 genotypes was three-fold higher, and the C_max value of omeprazole was three-fold higher compared with subjects with the same genotype in the single-dose study. However, there were few differences in the AUC and C_max of omeprazole between the *2/*3 genotype in the single-dose study and the homozygote for the CYP2C19*2 (*2/*2) in the repeated-dose study. Conclusion: Subjects with *1/*1, *1/*2 and *1/*3 genotypes in the repeated-dose study had lower CYP2C19 activity than subjects of the same genotype in the single-dose study. The difference in omeprazole metabolism between subjects with different genotypes observed on day 1 seemed to disappear after 7 days of repeated-dose administration. Received: 5 June 1998 / Accepted in revised form: 27 October 1998     

Toxicokinetic Study of Recombinant Human Heparin-Binding Epidermal Growth Factor-Like Growth Factor (rhHB-EGF) in Female Sprague Dawley Rats

Purpose To determine the toxicity and pharmacokinetics of recombinant heparin-binding epidermal growth factor-like growth factor in female Sprague Dawley rats following intra-bladder and intravenous administration. Materials and Methods rhHB-EGF was administered once daily for 6 or 27 days at doses of 3, 10, or 30 μg/kg. ¹²⁵I-rhHB-EGF was administered on day 7 or 28 for pharmacokinetic analysis. Toxicity was assessed by general appearance and behavior, gross necropsy, blood chemistry and microscopic evaluation. Results Plasma AUCss of [¹²⁵I] rhHB-EGF equivalents following IB administration for 7 days were 4.28 ± 2.29, 7.75 ± 2.70, and 7.11 ± 1.42 ng ml⁻¹ h⁻¹ at doses of 3, 10, and 30 μg/kg, respectively. Following IV administration, the AUCss on day 7 increased from 27.0 ± 2.66 to 124 ± 5.09 and 385.11 ± 7.57 ng ml⁻¹ h⁻¹ with increasing the dose from 3 to 10 and 30 μg/kg. Similar AUCss data was obtained after 28 day administration. No toxicity was evident upon gross examination. Histologic examination revealed subacute inflammation and lymphocytic infiltration of the urinary bladder in animals from all groups dosed by the IB route. Conclusions Plasma and bladder concentrations of recombinant human [¹²⁵I] rhHB-EGF equivalents were significantly lower following the IB route than following IV administration. Histologic tissue examination indicated no toxicity attributable to rhHB-EGF.     

Absence of interaction between erythromycin and a single dose of clozapine

Objective: To study the suggested pharmacokinetic interaction between erythromycin, a strong inhibitor of CYP3A4, and clozapine. Methods: Twelve healthy male volunteers received a single dose of 12.5 mg of clozapine alone or in combination with a daily dose of 1500 mg erythromycin in a randomised crossover study. Clozapine and its metabolites clozapine-N-oxide and desmethyl-clozapine were measured in serum samples which were collected during a 48 h period and in a sample of the urine secreted over the interval 0–12 h. Results: There were no significant differences in mean area under the serum concentration time curves (1348 (633) nmol h · 1⁻¹ in the control phase and 1180 (659) nmol h · 1⁻¹ in the erythromycin phase), terminal half-lives (19 (13) h and 15 (6) h, respectively), peak serum concentrations (92 (53) nmol · 1⁻¹ and 77 (40) nmol · 1⁻¹, respectively), time to peak serum concentrations (1.4 (0.7) h and 1.5 (1.0) h, respectively) or apparent oral clearances of clozapine (34 (15) l · h⁻¹ and 46 (37) l · h⁻¹, respectively). There were no significant differences in partial metabolic clearances to clozapine-N-oxide (5.1 (3.6) l · h⁻¹ and 7.8 (9.4) l · h⁻¹, respectively) or to desmethyl-clozapine (1.5 (1.3) l · h⁻¹ and 1.8 (1.7) l · h⁻¹, respectively) or in renal clearances of clozapine (0.8 (0.5) l · h⁻¹ and 1.0 (0.7) l · h⁻¹, respectively) between the two phases. Conclusion: These results demonstrate that erythromycin at a clinically relevant dosage does not inhibit the metabolism of clozapine. Hence, CYP3A4 seems to be of minor importance in the disposition of clozapine in humans at least when clozapine is taken at a low single dose. Received: 26 August 1998 / Accepted in revised form: 8 January 1999     

Comparative pharmacokinetics of dirithromycin and erythromycin in normal volunteers with special regard to accumulation in polymorphonuclear leukocytes and in saliva

Objective: In a randomized cross-over study, we assessed pharmacokinetics and intracellular concentrations in polymorphonuclear leukocytes (PMN) and saliva of erythromycin and erythromycylamine, the active metabolite of dirithromycin. Methods: Ten healthy volunteers received 1 g erythromycin b.i.d. or 500 mg dirithromycin qd for 5 days (wash out period, 35 days). Concentrations of erythromycin and erythromycylamine were measured in serum, urine, saliva, and granulocytes by bioassay and high-performance liquid chromatography (HPLC) on days 1, 3, and 5 of each study period, respectively. Results: While maximal serum concentrations (C_max) and the area under the data (AUD_tot) of erythromycin were significantly higher (C_max 1.44 mg · l⁻¹, AUD_tot 5.66 mg · h · l⁻¹) than those of erythromycylamine (C_max 0.29 mg · l⁻¹, AUD_tot 1.96 mg · h · l⁻¹), erythromycylamine had a significantly higher mean residence time (21 h) than erythromycin (5.5 h). Erythromycylamine accumulated significantly more in PMN than erythromycin;␣the accumulation factor of erythromycylamine was 100 with a maximal intracellular concentration of 13.4 mg · l⁻¹, whereas the maximal accumulation factor of erythromycin was 4 with a maximal intracellular concentration of 6.1 mg · l⁻¹. There were no significant differences in maximal saliva concentrations (erythromycin 0.35 mg · l⁻¹, erythromycylamine 0.31 mg · l⁻¹). Received: 16 September 1996 / Accepted in revised form: 12 February 1997     

Determination of the relative bioavailability of nedocromil sodium to the lung following inhalation using urinary excretion

Objective: To determine the effects of cimetidine on the steady-state pharmacokinetics and pharmacodynamics of atorvastatin, a 3-hydroxymethylglutaryl coenzyme A (HMG-CoA) reductase inhibitor. Methods: Twelve healthy subjects participated in a randomized two-way crossover study. Each subject received atorvastatin 10 mg every morning for 2 weeks and atorvastatin 10 mg every morning with cimetidine 300 mg four times a day for 2 weeks, separated by a 4-week washout period. Steady-state pharmacokinetic parameters (based on an enzyme inhibition assay) and lipid responses were compared. Results: Pharmacokinetic parameters and lipid responses were similar following administration of atorvastatin alone and atorvastatin with cimetidine. Mean values for C_max (the maximum concentration) were 5.11 ng · eq · ml⁻¹ and 4.54 ng eq · ml⁻¹, for t_max (the time to reach maximum concentration) 2.2 h and 1.3 h, for AUC_0–24 (area under the concentration-time curve from time 0 h to 24 h) 58.6 ng eq · h · ml⁻¹ and 58.5 ng eq · h · ml⁻¹, and for t_1/2 (terminal half-life) 10.1 h and 17.0 h, respectively, following administration of atorvastatin alone and atorvastatin with cimetidine. Following treatment with atorvastatin alone and atorvastatin with cimetidine, mean values for the percentage change from baseline for total cholesterol were −29.5% and −29.9%, for low-density lipoprotein (LDL) cholesterol −41.0% and −42.6%, for high-density lipoprotein (HDL) cholesterol 6.3% and 5.8%, and for triglycerides −33.8% and −25.8%, respectively. Conclusions: The rate and extent of atorvastatin absorption and the effects of atorvastatin on LDL-cholesterol responses are not influenced by coadministration of cimetidine. Received: 17 February 1997 / Accepted in revised form: 3 November 1997