Discovery of AMG 853, a CRTH2 and DP Dual Antagonist

相似文献   

Application of Lipid Microspheres to Prepare a Thromboxane A2 Receptor Antagonist Aerosol for Inhalation

Abstract

The methyl ester of a new thromboxane A₂ receptor antagonist, (+)S-145, i.e.(1R,2S,3S,4S)-(5Z)-7-(3-phenylsulfonyl-aminobicyclo[2,2,1]hept-2-yl)heptenoic acid, was incorporated into lipid microspheres (lipo S-145-Me) and its pharmacological effect and tissue distribution were examined in guinea pigs following aerosol delivery. Bronchoconstrictive responses induced by intravenous injection of U46619 or the inhalation of ovalbumin were suppressed in a dose-dependent manner by aerosol inhalation of lipo S-145-Me, which was 3-10 times more potent that the unencapsulated calcium dihydrate of the original drug (S-1452). There was no significant difference in the airway tissue distribution of labelled lipo S-145-Me versus S-1452 after 2 or 5 min of inhalation, but the encapsulated drug showed marked accumulation in the lungs after 30 min of inhalation. The in vitro uptake of lipo [¹⁴C] S-145-Me by fresh human neutrophils and an eosinophil cell line was respectively 7 times and 3.5 times higher than that of [¹⁴C] S-1452. These results suggest that lipo S-145-Me has the potential to be used as an inhalational antiasthma agent, and that its effect may be partly attributable to a for inflammatory cells which are responsible for allergic airway inflammation.     

Neurogenesis-Independent Antidepressant-Like Effects on Behavior and Stress Axis Response of a Dual Orexin Receptor Antagonist in a Rodent Model of Depression

Growing evidence indicates that an increase of orexin (or hypocretin) signaling is involved in the pathophysiology of major depression, but little is known regarding the causal link between the orexinergic system and depressive-like states. Here we blocked orexin receptors in mice subjected to unpredictable chronic mild stress (UCMS) to investigate putative antidepressant-like effects of this treatment, as well as the underlying mechanisms. BALB/c mice were exposed to 9 weeks of UCMS and from the third week onward treated daily with fluoxetine (20 mg/kg per day, per os) or with the dual orexin receptor antagonist almorexant (100 mg/kg per day, per os). The effects of UCMS regimen and pharmacological treatments were assessed by physical measures and behavioral testing. The dexamethasone suppression test was performed to examine the integrity of the negative feedback of the hypothalamic-pituitary-adrenal (HPA) axis, and immunohistochemical markers were used to assess cell proliferation (Ki-67), immature newborn neurons (doublecortin), and mature newborn neurons (5-bromo-2′-deoxyuridine/NeuN) in the dorsal and ventral parts of the hippocampus. Our results show that 7 weeks of fluoxetine or almorexant treatments counteract the UCMS-induced physical and behavioral alterations. Both treatments prevented the HPA axis dysregulation caused by UCMS, but only fluoxetine reversed the UCMS-induced decrease of hippocampal cell proliferation and neurogenesis, while chronic almorexant treatment decreased cell proliferation and neurogenesis specifically in the ventral hippocampus. Taken together, this is the first evidence that pharmacological blockade of the orexinergic system induces a robust antidepressant-like effect and the restoration of stress-related HPA axis defect independently from a neurogenic action.     

Evaluation of the Safety,Tolerability, and Pharmacokinetic Profiles of TP0473292 (TS-161), A Prodrug of a Novel Orthosteric mGlu2/3 Receptor Antagonist TP0178894, in Healthy Subjects and Its Antidepressant-Like Effects in Rodents

BackgroundTP0473292 (the active ingredient of TS-161) is a prodrug of a novel metabotropic glutamate (mGlu) 2/3 receptor antagonist being developed for the treatment of patients with depression. This study evaluated the safety, tolerability, and pharmacokinetics of orally administered TS-161 in healthy subjects.MethodsThis was a first-in-human, phase 1, randomized, double-blind, placebo-controlled, single-ascending dose (15–400 mg TS-161) and 10-day multiple-ascending dose (50–150 mg TS-161) study in healthy subjects, conducted from June 2019 through February 2020. Plasma and urine concentrations of the prodrug and its metabolites, and cerebrospinal fluid (CSF) concentrations of the active metabolite TP0178894 were measured to evaluate the pharmacokinetic profiles after oral administration of TS-161.ResultsFollowing single and multiple doses, TP0473292 was extensively converted into its active metabolite TP0178894. Plasma concentrations of TP0178894 reached peak (C_max) within 5 hours post dose and declined with a t_1/2 <13 hours. Plasma exposures of TP0178894 increased with increasing dose. TP0178894 penetrated into CSF and reached a C_max of 9.892 ng/mL at a single dose of 100 mg, which was comparable with IC₅₀ values of antagonist activity at mGlu2/3 receptors. The most frequently observed adverse events that showed exposure-related incidence during the study were nausea, vomiting, and dizziness.ConclusionsThe mGlu2/3 receptor antagonist prodrug TP0473292 is safe and well-tolerated, is orally bioavailable in humans with extensive conversion into the active metabolite TP0178894 with sufficient CSF penetration to exert the anticipated pharmacological effects, and is a promising candidate for further clinical development in treatment of patients with depression.     

Discovery of TAK-925 as a Potent,Selective, and Brain-Penetrant Orexin 2 Receptor Agonist

TAK-925, a potent, selective, and brain-penetrant orexin 2 receptor (OX2R) agonist, [methyl (2R,3S)-3-((methylsulfonyl)amino)-2-(((cis-4-phenylcyclohexyl)oxy)methyl)piperidine-1-carboxylate, 16], was identified through the optimization of compound 2, which was discovered by a high throughput screening (HTS) campaign. Subcutaneous administration of compound 16 produced wake-promoting effects in mice during the sleep phase. Compound 16 (TAK-925) is being developed for the treatment of narcolepsy and other related disorders.     

Betulinic Acid Ameliorates the T-2 Toxin-Triggered Intestinal Impairment in Mice by Inhibiting Inflammation and Mucosal Barrier Dysfunction through the NF-κB Signaling Pathway

T-2 toxin, a trichothecene mycotoxin produced by Fusarium, is widely distributed in crops and animal feed and frequently induces intestinal damage. Betulinic acid (BA), a plant-derived pentacyclic lupane-type triterpene, possesses potential immunomodulatory, antioxidant and anti-inflammatory biological properties. The current study aimed to explore the protective effect and molecular mechanisms of BA on intestinal mucosal impairment provoked by acute exposure to T-2 toxin. Mice were intragastrically administered BA (0.25, 0.5, or 1 mg/kg) daily for 2 weeks and then injected intraperitoneally with T-2 toxin (4 mg/kg) once to induce an intestinal impairment. BA pretreatment inhibited the loss of antioxidant capacity in the intestine of T-2 toxin-treated mice by elevating the levels of CAT, GSH-PX and GSH and reducing the accumulation of MDA. In addition, BA pretreatment alleviated the T-2 toxin-triggered intestinal immune barrier dysregulation by increasing the SIgA level in the intestine at dosages of 0.5 and 1 mg/kg, increasing IgG and IgM levels in serum at dosages of 0.5 and 1 mg/kg and restoring the intestinal C3 and C4 levels at a dosage of 1 mg/kg. BA administration at a dosage of 1 mg/kg also improved the intestinal chemical barrier by decreasing the serum level of DAO. Moreover, BA pretreatment improved the intestinal physical barrier via boosting the expression of ZO-1 and Occludin mRNAs and restoring the morphology of intestinal villi that was altered by T-2 toxin. Furthermore, treatment with 1 mg/kg BA downregulated the expression of p-NF-κB and p-IκB-α proteins in the intestine, while all doses of BA suppressed the pro-inflammatory cytokines expression of IL-1β, IL-6 and TNF-α mRNAs and increased the anti-inflammatory cytokine expression of IL-10 mRNA in the intestine of T-2 toxin-exposed mice. BA was proposed to exert a protective effect on intestinal mucosal disruption in T-2 toxin-stimulated mice by enhancing the intestinal antioxidant capacity, inhibiting the secretion of inflammatory cytokines and repairing intestinal mucosal barrier functions, which may be associated with BA-mediated inhibition of the NF-κB signaling pathway activation.     

Ascorbic Acid for the Safe Use of a Sunscreen Agent: Accumulation of Nano Zinc Oxide and Titanium Dioxide on the Skin

Objective

Physical UV absorbers such as titanium dioxide or zinc oxide have been found to be highly protective against ultraviolet radiation. Sun protection factor depends on the accumulation of the minerals on the skin. UV-absorbing agents must accumulate within the upper skin layers in order to provide a dense light-absorbing layer and guarantee water resistance. The aim of this work was to increase the skin deposition and efficacy of sunscreens without increasing their skin permeation. The application possibility of EDX to determine the quantitative elemental composition of zinc and titanium on the skin surface was studied.

Method

The changes induced in the skin deposition of physical UV absorbers in conjunction with ascorbic acid were studied. In vitro skin permeation and X-ray elemental analysis were carried out to determine the mineral skin deposition effect of ascorbic acid.

Key findings

Results indicate that ascorbic acid may significantly increase the skin deposition (p < 0.05) of these minerals on the skin without increasing their skin permeation (p > 0.05). Flow through diffusion cell and X-ray elemental analyses appear to be complementary and show that ascorbic acid is able to increase accumulation of sunscreen on the skin.     

Effect of food intake on pharmacokinetics and effects of a new thromboxane A2 receptor antagonist,S-1452

Objective: To examine the effect of food ingestion on the pharmacokinetics of a new thromboxane A₂ (TXA₂) receptor antagonist, S-1452, and the inhibitory effect on platelet aggregation. Methods: Fifty milligrams of S-1452 was given orally to eight healthy subjects with or without food. Blood samples for determinations of plasma drug concentrations and of its effects on platelet aggregation were taken for a 12-h post-drug period. Results: The maximum plasma concentration of S-1452 was reduced by 47% and the time to maximum concentration was prolonged from 0.5 to 1.9 h after dosing with food. The inhibitory effect of S-1452 on platelet aggregations induced by U-46619, a TXA₂ receptor agonist, and collagen persisted up to 9 h after dosing with and without food. The degrees of inhibition in the two trials did not differ significantly at any point. Conclusion: These results suggest that although the absorption of S-1452 is delayed and, consequently, its plasma concentration is decreased after dosing with food, the inhibitory effect on platelet aggregation is not significantly influenced after 50 mg of the drug. Received: 1 August 1995/Accepted in revised form: 24 October 1995     

Transcellular Transport of Benzole Acid Across Caco-2 Cells by a pH-Dependent and Carrier-Mediated Transport Mechanism

The pH-dependent transcellular transport of [¹⁴ C]benzoic acid across a Caco-2 cell monolayer is shown to be mediated by a monocarboxylic acid-specific carrier-mediated transport system, localized on the apical membrane. Evidence for the carrier-mediated transport of benzoic acid includes (a) the significant temperature and concentration dependence, (b) the metabolic energy dependence, (c) the inhibition by unlabeled benzoic acid and other monocarboxylic acids, (d) countertransport effects on the uptake of [¹⁴C]benzoic acid, and (e) effects of a proteinase (papain) and amino acid-modifying reagents. Furthermore, since carbonylcyanide p-trifluoromethoxyphenylhydrazone and nigericin significantly inhibited the transport of [¹⁴C] benzoic acid, the direct driving force for benzoic acid transport is suggested to be the inwardly directed proton gradient. From these results, together with previous observations using intestinal brush border membrane vesicles, the pH dependence of the transcellular transport of certain organic weak acids across Caco-2 cells is considered to result mainly from a proton gradient-dependent, carrier-mediated transport mechanism, rather than passive diffusion according to the pH-partition theory.     

Effect of netazepide,a gastrin/CCK2 receptor antagonist,on gastric acid secretion and rabeprazoleinduced hypergastrinaemia in healthy subjects

Aims

To compare gastric acid suppression by netazepide, a gastrin/CCK₂ receptor antagonist, with that by a proton pump inhibitor (PPI), and to determine if netazepide can prevent the trophic effects of PPI-induced hypergastrinaemia.

Methods

Thirty healthy subjects completed a double-blind, randomized, parallel group trial of oral netazepide and rabeprazole, alone and combined, once daily for 6 weeks. Primary end points were: basal and pentagastrin-stimulated gastric acid and 24 h circulating gastrin and chromogranin A (CgA) at baseline, start and end of treatment, gastric biopsies at baseline and end of treatment and basal and pentagastrin-stimulated gastric acid and dyspepsia questionnaire after treatment withdrawal.

Results

All treatments similarly inhibited pentagastrin-stimulated gastric acid secretion. All treatments increased serum gastrin, but the combination and rabeprazole did so more than netazepide alone. The combination also reduced basal acid secretion.Rabeprazole increased plasma CgA, whereas netazepide and the combination reduced it. None of the biopsies showed enterochromaffin-like (ECL) cell hyperplasia. Withdrawal of treatments led neither to rebound hyperacidity nor dyspepsia.

Conclusions

Netazepide suppressed pentagastrin-stimulated gastric acid secretion as effectively as did rabeprazole. The reduction in basal acid secretion and greater increase in serum gastrin by the combination is consistent with more effective acid suppression. Despite our failure to show rabeprazole-induced ECL cell hyperplasia and rebound hyperacidity, the increase in plasma CgA after rabeprazole is consistent with a trophic effect on ECL cells, which netazepide prevented. Thus, netazepide is a potential treatment for the trophic effects of hypergastrinaemia and, with or without a PPI, is a potential treatment for acid-related conditions.     

Comparative Analysis of Percutaneous Absorption Enhancement by d-Limonene and Oleic Acid Based on a Skin Diffusion Model

Percutaneous absorption-enhancing effects of d-limonene and oleic acid were investigated using three model drugs with different lipophilicities in in vitro diffusion experiments with guinea pig skin. Pretreatment of the skin with d-limonene resulted in a large penetration enhancement for the lipophilic butylparaben (BP) and amphiphilic 6-mercaptopurine (6-MP) but had little effect on the hydrophilic mannitol (MT). Oleic acid caused a large effect only on 6-MP penetration. The penetration profiles were analyzed with a two-layer skin diffusion model consisting of stratum corneum with polar and nonpolar routes and viable epidermis plus dermis. Through curve-fitting, six parameters corresponding to drug diffusivity and partitioning in these three regions of the skin were obtained, and the mechanisms of enhancers were assessed in comparison with those of l-geranylazacycloheptan-2-one (GACH) reported previously. Increased penetration was caused mainly by modification of the barrier property of the nonpolar route in the stratum corneum in all cases. In the nonpolar route, d-limonene increased mainly drug diffusivity, while GACH enhanced predominately drug partitioning. On the other hand, oleic acid moderately increased both parameters.     

丹酚酸B对H_2O_2引起H9c2细胞氧化损伤的保护作用及可能靶点蛋白寻找

观察丹酚酸B对H2O2引起H9c2细胞氧化损伤的影响,在分子水平上探讨丹酚酸B对H2O2诱导氧化损伤保护作用的可能机制。用H2O2刺激H9c2建立体外心肌细胞氧化损伤模型,MTT测定心肌细胞的存活率,Hochest33342染色观察细胞凋亡,并用蛋白质组学技术寻找丹酚酸B在心肌细胞中的可能蛋白质靶点。丹酚酸B可以提高心肌细胞在H2O2处理下的存活率,减少H2O2导致的细胞凋亡,丹酚酸B预处理组与单用H2O2组相比有8个差异表达蛋白,其中Heat shock 27 kua protein蛋白下调,cytochrome C oxidase、myo-sinlight polypeptide 6、myosin、myosin regulatory light chain 2、myosin regulatory light chain 9、tropomyosin和secreted acidic cysteine rich glycoprotein蛋白上调,差异表达的蛋白可能在丹酚酸B的保护机制中发挥重要作用。     

Effect of the H2-Receptor Antagonist Cimetidine,on the Pharmacokinetics and Pharmacodynamics of the H1-Receptor Antagonists Hydroxyzine and Cetirizine in Rabbits

The effects of coadministration of the H₂-receptor antagonist cimetidine on the pharmacokinetics and pharmacodynamics of the H₁ receptor antagonists hydroxyzine and cetirizine were studied in rabbits. A single dose of hydroxyzine, 10 mg (Experiment A), or cetirizine, 10 mg (Experiment B), was given intravenously on three occasions: 2 weeks before cimetidine administration, after cimetidine, 100 mg/kg, had been given every 12 hr for 1 week, and 2 weeks after the cimetidine was discontinued. Serum concentrations of hydroxyzine and cetirizine, the active metabolite of hydroxyzine arising in vivo (Experiment A), or cetirizine (Experiment B) were measured by HPLC. The pharmacologic effects of hydroxyzine and cetirizine were monitored by measuring the suppression of histamine-induced wheals, using an IBM-PC and digitizer. The hydroxyzine and cetrizine half-life and AUG₀ values were significantly increased and the systemic clearance rates were significantly decreased in the presence of cimetidine. Similar results were obtained when cetirizine was administered de novo. Wheal suppression produced by hydroxyzine or cetirizine was increased and prolonged in the presence of cimetidine. The synergism observed between hydroxyzine or cetirizine and cimetidine in suppression of the histamine-induced cutaneous response may be due to a pharmacokinetic interaction.     

Synthesis and in vivo evaluation in mice of [123I]‐(4‐fluorophenyl)[1‐(3‐iodophenethyl)piperidin‐4‐yl]methanone as a potential SPECT‐tracer for the serotonin 5‐HT2A receptor

This work reports the synthesis, radiolabelling and in vivo evaluation in NMRI mice of [¹²³I]‐(4‐fluorophenyl)[1‐(3‐iodophenethyl)piperidin‐4‐yl]methanone ([¹²³I]‐3‐I‐CO) as a potential SPECT tracer for the 5‐HT_2A receptor. The tributylstannylprecursor was synthesized with a 15% overall yield. Radiolabelling was performed using an electrophilic iododestannylation with yields of 85%. Radiochemical purity was always >95%. Log P was determined to be 3.10±0.10. The tracer showed good uptake in mouse brain (6.3±1.3% ID/g tissue at 10 min p.i., 2±0.36% ID/g tissue at 1 h p.i.). These results warrant further research in larger animals to determine suitability of [¹²³I]‐3‐I‐CO as a 5‐HT_2A tracer. Copyright © 2007 John Wiley & Sons, Ltd.     

超临界萃取当归挥发油及阿魏酸正交试验研究

目的：优选超临界二氧化碳流体（SFE-CO2）萃取当归挥发油和阿魏酸的工艺条件。方法：以阿魏酸的提取量和挥发油的得率为考察指标，以高效液相色谱法测定阿魏酸含量，采用正交试验对提取条件优选。结果：优选萃取条件是：当归粉碎为20—60目，75％的乙醇作为夹带剂，萃取压力30Mpa，萃取温度60℃。结论：优选工艺萃取的挥发油和阿魏酸提取量较高，所得浸膏量较低。     

Impact of CYP2C19 phenotypes on escitalopram metabolism and an evaluation of pupillometry as a serotonergic biomarker

Purpose  To investigate the impact of cytochrome P450 2C19 (CYP2C19) phenotypes on escitalopram metabolism and to evaluate pupillometry as a serotonergic biomarker. Methods  This was a double-blind, crossover design study with single and multiple doses of 10 mg escitalopram and placebo in panels of CYP2C19 extensive (EM) and poor metabolisers (PM). Pupillometry was measured by a NeurOptics Pupillometer-PLR. Results  Five PM and eight EM completed the study. The CYP2C19 phenotype significantly affected the metabolism of escitalopram. The area under the time–plasma concentration curve (AUC_0-24) was 1.8-fold higher in PM than in EM after both single and multiple doses. Escitalopram treatment did not affect the maximum pupil size, but it did statistically significantly decrease the relative amplitude of the pupil light reflex compared to the placebo; this effect was equal in both phenotype groups. Conclusions  The CYP2C19 polymorphism affects escitalopram metabolism, but the difference does not justify dose adjustment. The puzzling results from pupillometry can be due to interplay between a central and a local serotonergic effect. Based on these results, pupillometry can not be recommended as a serotonergic biomarker.     

Tachykinin NK2 receptors: characterization in the rat small intestine by the use of a peptide agonist and a nonpeptide antagonist as radioligands

The tachykinin NK₂ receptors present in membranes of rat small intestine were characterized by means of tachykinin receptor agonists and antagonists, by using the natural agonist, NKA, or the nonpeptide antagonist SR 48968, as radioligands. The affinity of the antagonists was independent of the radioligand used, whereas the NK₂ receptor selective agonists showed a different binding profile according to the radioligand. In particular, when using [¹²⁵I]NKA, NKA and NKA_(4–10) bound to a high affinity site, whilst [β-Ala⁸]NKA_(4–10) and GR 64349 bound to a high and a low affinity site; the high affinity site was still detected in the presence of 100μM GppNHp which produced a strong inhibition of the specific binding of [¹²⁵I]NKA. On the other hand, when using [³H]SR 48968, NKA bound to a high affinity site, [β-Ala⁸]NKA_(4–10) bound to a low affinity site and NKA_(4–10) and GR 64349 bound to a high and a low affinity site; in this case, the high affinity site was no longer detected in the presence of 1μM GppNHp, which did not reduce the specific binding of [³H]SR 48968 to NK₂ receptors. We interpreted these data as an indication that in the rat small intestine membranes [¹²⁵I]NKA labels multiple conformations of G protein-coupled NK₂ receptor which are distinguished by the use of selective receptor agonists, but not by peptide or nonpeptide antagonists. Received: 11 September 1996 / Accepted: 12 March 1997     

人尿中对乙酰氨基苯甲酸和N,N-二甲氨基-2-丙醇的液质联用法测定及尿药排泄特征研究

目的：建立分别测定人尿中对乙酰氨基苯甲酸和N,N-二甲氨基-2-丙醇的液质联用法,考察二者在中国健康受试者尿液中的排泄特征。方法：以LC-MS法测定尿样中对乙酰氨基苯甲酸,色谱采用Amethyst C18柱（150 mm×2.1 mm,5μm）,流动相为甲醇-0.1%甲酸水溶液（25∶75）,流速为0.4 mL.min-1;质谱采用气动辅助电喷雾离子化和正离子选择性离子检测。以LC-MS/MS法测定尿样中N,N-二甲氨基-2-丙醇,色谱采用Hedera CN柱（150 mm×2.1 mm,5μm）,流动相为乙腈-5 mmol.L-1醋酸铵水溶液（含0.03%甲酸）（55∶45）,流速为0.35 mL.min-1;质谱采用气动辅助电喷雾离子化和正离子多反应检测。10名受试者单次口服异丙肌苷片（1.0 g）,测定对乙酰氨基苯甲酸和N,N-二甲氨基-2-丙醇的尿药排泄参数。结果：对乙酰氨基苯甲酸的尿药浓度在0.202 0～202.0 mg.L-1范围内线性关系良好,平均回收率大于97.7%;N,N-二甲氨基-2-丙醇尿药浓度在0.797 8～398.9 mg.L-1范围内线性关系良好,平均回收率大于99.1%。受试者服药后,对乙酰氨基苯甲酸和N,N-二甲氨基-2-丙醇分别在6和12 h后基本随尿排泄完全,36 h内其平均尿药累积排泄百分率分别为（30.7±5.7）%和（49.0±8.6）%。结论：本法适用于人尿中对乙酰氨基苯甲酸和N,N-二甲氨基-2-丙醇的测定及其尿药排泄特征研究。     

ATP-Dependent Transport of a Novel Thromboxane A2 Receptor Antagonist, [2-(4-Chlorophenylsulfonylaminomethyl)-Indan-5-yl]Acetate (Z-335) and Its Xenobiotic Taurine Conjugate (Z-335-Tau) by Rat Bile Canalicular Membrane Vesicles

PURPOSE: The characteristics of bile canalicular transport processes for xenobiotic taurine conjugates have not yet been clarified. To elucidate the biliary excretion characteristics of xenobiotic taurine conjugates, we investigated the transport of a novel thromboxane A2 receptor antagonist, Z-335, and its taurine conjugate (Z-335-Tau) across the bile canalicular membrane. METHODS: We examined the uptake of Z-335 and Z-335-Tau by isolated bile canalicular membrane vesicles (CMVs) from Sprague Dawley and Eisai-hyperbilirubinemic rats (EHBRs) which EHBRs have a hereditary defect of canalicular multidrug resistance-associated protein 2 (Mrp2) function. Also, the in vitro and in vivo kinetics of Z-335-Tau uptake and excretion were compared. RESULTS: Z-335 uptake by CMVs from normal rats exhibited marked ATP-dependence, whereas ATP-dependent uptake of Z-335 into CMVs from EHBRs was not observed. In contrast, Z-335-Tau uptake into CMVs from both normal rats and EHBRs was ATP dependent. The initial uptake velocity was concentration-dependent, with an in vitro Michaelis constant for initial uptake of 189 microM, which was similar to the in vivo value. CONCLUSIONS: The biliary excretion of Z-335 involves Mrp2, whereas that of Z-335-Tau involves active transport systems that remain intact in EHBRs and show marked ATP dependence, which ATP-dependent transport is involved in the biliary excretion of Z-335-Tau in vivo.     

Impact of the Inoculation Method of Geotrichum candidum,Used as Biocontrol Agent,on T-2 Toxin Produced by Fusarium sporotrichioides and F. langsethiae during the Malting Process

In malt production, steeping and germination steps offer favorable environmental conditions for fungal proliferation when barley is already contaminated by Fusarium species, T-2 toxin producers. However, the use of G. candidum as a biocontrol agent can prevent this proliferation. Indeed, in previous work, a correlation between phenyllactic acid (PLA) production by G. candidum and the reduction in Fusarium sporotrichioides and F. langsethiae growth and T-2 toxin concentration was demonstrated. In the present study, to improve the efficiency of G. candidum, the effects of the inoculum concentration and the inoculation method of G. candidum on PLA and T-2 toxin concentrations were evaluated. First, co-culture experiments with Fusarium species and G. candidum were conducted in a liquid synthetic medium. The results showed that inoculation of G. candidum in the freeze-dried form at 0.4 g/L allowed the production of PLA from the second day of incubation associated with a reduction in T-2 toxin concentration of 82% and 69% produced by F. sporotrichioides and F. langsethiae, respectively. Moreover, the activated form of G. candidum at 0.4 g/L enhanced PLA concentration leading to better T-2 toxin reduction. Second, experiments were conducted on artificially infected barley kernels with both Fusarium species under conditions mimicking the malting step. As for co-culture experiments, the use of the activated form of G. candidum was established as the best condition for T-2 toxin concentration reduction for a 3 day malting period.