Sustained oxidation of Tea-Fe(III)/H2O2 simultaneously achieves sludge reduction and carbamazepine removal: The crucial role of EPS regulation.

Establishing an economic and sustained Fenton oxidation system to enhance sludge dewaterability and carbamazepine (CBZ) removal rate is a crucial path to simultaneously achieve sludge reduction and harmless. Leveraging the principles akin to "tea making", we harnessed tea waste to continually release tea polyphenols (TP), thus effectively maintaining high level of oxidation efficiency through the sustained Fenton reaction. The results illustrated that the incorporation of tea waste yielded more favorable outcomes in terms of water content reduction and CBZ removal compared to direct TP addition within the Fe(III)/hydrogen peroxide (H2O2) system. Concomitantly, this process mainly generated hydroxyl radical (•OH) via three oxidation pathways, effectively altering the properties of extracellular polymeric substances (EPS) and promoting the degradation of CBZ from the sludge mixture. The interval addition of Fe(III) and H2O2 heightened extracellular oxidation efficacy, promoting the desorption and removal of CBZ. The degradation of EPS prompted the transformation of bound water to free water, while the formation of larger channels drove the discharge of water. This work achieved the concept of treating waste with waste through using tea waste to treat sludge, meanwhile, can provide ideas for subsequent sludge harmless disposal.

Pharmacokinetic Drug-Drug Interactions among Antiepileptic Drugs, Including CBD, Drugs Used to Treat COVID-19 and Nutrients.

Anti-epileptic drugs (AEDs) are an important group of drugs of several generations, ranging from the oldest phenobarbital (1912) to the most recent cenobamate (2019). Cannabidiol (CBD) is increasingly used to treat epilepsy. The outbreak of the SARS-CoV-2 pandemic in 2019 created new challenges in the effective treatment of epilepsy in COVID-19 patients. The purpose of this review is to present data from the last few years on drug-drug interactions among of AEDs, as well as AEDs with other drugs, nutrients and food. Literature data was collected mainly in PubMed, as well as google base. The most important pharmacokinetic parameters of the chosen 29 AEDs, mechanism of action and clinical application, as well as their biotransformation, are presented. We pay a special attention to the new potential interactions of the applied first-generation AEDs (carbamazepine, oxcarbazepine, phenytoin, phenobarbital and primidone), on decreased concentration of some medications (atazanavir and remdesivir), or their compositions (darunavir/cobicistat and lopinavir/ritonavir) used in the treatment of COVID-19 patients. CBD interactions with AEDs are clearly defined. In addition, nutrients, as well as diet, cause changes in pharmacokinetics of some AEDs. The understanding of the pharmacokinetic interactions of the AEDs seems to be important in effective management of epilepsy.

Fabrication of Carbamazepine Cocrystals: Characterization, In Vitro and Comparative In Vivo Evaluation.

Carbamazepine (CBZ) is an antiepileptic drug having low bioavailability due to its hydrophobic nature. In the current study, efforts are made to investigate the effect of dicarboxylic acid coformer spacer groups (aliphatic chain length) on physicochemical properties, relative humidity (RH) stability, and oral bioavailability of CBZ cocrystals. Slurry crystallization technique was employed for the preparation of CBZ cocrystals with the following coformers: adipic (AA), glutaric (GA), succinic (SA), and malonic acid (MA). Powder X-ray diffractometry and Fourier-transform infrared spectroscopy confirmed cocrystal preparation. Physicochemical properties, RH stability, and oral bioavailability of cocrystals were investigated. Among the prepared cocrystals, CBZ-GA showed maximum solubility as well as improved dissolution profile (CBZ-GA > CBZ-MA > CBZ-AA > pure CBZ > CBZ-SA) in ethanol. Maximum RH stability was shown by CBZ-AA, CBZ-SA, and CBZ-MA. In vivo studies confirmed boosted oral bioavailability of cocrystals compared to pure CBZ. Furthermore, in vivo studies depicted the oral bioavailability order of cocrystals as CBZ-GA > CBZ-MA > Tegral® > CBZ-AA > CBZ-SA > pure CBZ. Thus, pharmaceutical scientists can effectively employ cocrystallization technique for tuning physicochemical properties of hydrophobic drugs to achieve the desired oral bioavailability. Overall, results reflect no consistent effect of spacer group on physicochemical properties, RH stability, and oral bioavailability of cocrystals.

Environmentally friendly synthesized and magnetically recoverable designed ferrite photo-catalysts for wastewater treatment applications.

Fenton processes are promising wastewater treatment alternatives for bio-recalcitrant compounds. Three different methods (i.e., reverse microemulsion, sol-gel, and combustion) were designed to synthesize environmentally friendly ferrites as magnetically recoverable catalysts to be applied for the decomposition of two pharmaceuticals (ciprofloxacin and carbamazepine) that are frequently detected in water bodies. The catalysts were used in a heterogeneous solar photo-Fenton treatment to save the cost of applying high-energy UV radiation sources, and was performed under a slightly basic pH to avoid metal leaching and adding salts for pH adjustment. All the developed catalysts resulted in the effective treatment of ciprofloxacin and carbamazepine in both synthetic and real domestic wastewater. In particular, the sol-gel synthesized ferrite was more magnetic and more suitable for reuse. The degradation pathways of both compounds were elucidated for this treatment. The degradation of ciprofloxacin involved attacks to the quinolone and piperazine rings. The degradation pathway of carbamazepine involved the formation of hydroxyl carbamazepine and dihydroxy carbamazepine before yielding acridine by hydrogen abstraction, decarboxylation, and amine cleavage, which would be further oxidized.

Tuning down the environmental interests of organoclays for emerging pollutants: Pharmaceuticals in presence of electrolytes.

The impact of electrolytes on the adsorption of emerging pollutants: pharmaceuticals onto layered materials: a raw clay mineral and its nonionic and cationic organoclay derivatives was studied. The selected pharmaceuticals: amoxicillin, norfloxacin, sulfamethoxazole, metoprolol, carbamazepine, and trimethoprim show different electric charges: zwitterionic, anionic, cationic and neutral and hydrophobic character (different LogP). Without any salts, the set of complementary data obtained by UV and infrared spectroscopies, X-ray diffraction points out the importance of the electric charge which represents a key parameter in both the spontaneity and feasibility of the adsorption. In contrast, the hydrophobicity of the analytes plays a minor role but determines the magnitude of the adsorbed amount of pharmaceuticals onto organoclays. With a dual hydrophilic and hydrophobic behavior, nonionic organoclay appears to be the most polyvalent material for the removal of the pharmaceuticals. In the presence of electrolytes (NaCl at a concentration of 1 × 10-2 mol L-1), both nonionic and cationic organoclays show a decrease of their efficiencies, whereas the adsorption is particularly enhanced for Na-Mt except for the cationic species (trimethoprim and metoprolol). Thus, in realistic experimental conditions close to those of natural effluents, raw clay mineral appears as the most appropriate sorbent for the studied pharmaceuticals while it raises the question of the usefulness of organoclays in water remediation strategy.

Polygala tenuifolia-Acori tatarinowii herbal pair as an inspiration for substituted cinnamic α-asaronol esters: Design, synthesis, anticonvulsant activity, and inhibition of lactate dehydrogenase study.

Inspired by the traditional Chinese herbal pair of Polygala tenuifolia-Acori Tatarinowii for treating epilepsy, 33 novel substituted cinnamic α-asaronol esters and analogues were designed by Combination of Traditional Chinese Medicine Molecular Chemistry (CTCMMC) strategy, synthesized and tested systematically not only for anticonvulsant activity in three mouse models but also for LDH inhibitory activity. Thereinto, 68-70 and 75 displayed excellent and broad spectra of anticonvulsant activities with modest ability in preventing neuropathic pain, as well as low neurotoxicity. The protective indices of these four compounds compared favorably with stiripentol, lacosamide, carbamazepine and valproic acid. 68-70 exhibited good LDH1 and LDH5 inhibitory activities with noncompetitive inhibition type, and were more potent than stiripentol. Notably, 70, as a representative agent, was also shown as a moderately positive allosteric modulator at human α1ß2γ2 GABAA receptors (EC50 46.3 ±â€¯7.3 µM). Thus, 68-70 were promising candidates for developing into anti-epileptic drugs, especially for treatment of refractory epilepsies such as Dravet syndrome.

Alterations in Pharmacokinetics of Orally Administered Carbamazepine in Rats Treated with Sodium alginate: Possible Interaction between Therapeutic Drugs and Semi-solid Enteral Nutrients.

OBJECTIVE: The use of enteral nutrients plays an extremely important role in accurate nutrition management. Sodium alginate (SA) is frequently used for the semi-solidification of enteral nutrients. In the present study, we investigated whether the pharmacokinetic profile of orally administered carbamazepine (CBZ) is altered by a treatment with SA immediately before and after dosing of the drug. Furthermore, the adsorption effects of SA on CBZ were examined using an in vitro analysis. METHOD: SA was orally administered to rats just before and immediately after CBZ dosing. The CBZ concentration profile following its oral administration was analyzed by a non-compartmental method. The adsorption of CBZ onto SA was evaluated after centrifugation using an ultrafiltration device. FINDINGS: The serum concentration of orally administered CBZ at each sampling point was reduced by the treatment with SA, and the extent of the decrease observed in the concentration of CBZ was larger when SA was ingested immediately after administration of the drug, which was consistent with the alteration observed in the value of the area under the curve (AUC). No significant differences were noted in the elimination rate at the terminal phase (ke) among the groups. In the in vitro assay, CBZ was adsorbed by SA in the solution used to reflect fluid in the intestinal tract. CONCLUSIONS: The pharmacological efficacies of CBZ might be reduced by SA through the pharmacokinetic interactions, and that the careful attention should be paid to the timing of administration of CBZ and semi-solid enteral nutrients.

A four-year simulation of soil aquifer treatment using columns filled with San Gabriel Valley sand.

Two column pairs filled with 3.05-m of a sandy soil from the Upper San Gabriel Valley were operated for a period of four and ½ years on municipal effluent from the San Jose Creek Water Reclamation Plant operated by the Sanitation Districts of Los Angeles County (LACSD). One column pair was fed filtered, chlorinated effluent (tertiary effluent) for the entire period. The other pair was fed ozonated secondary effluent for 8-mo, ozonated secondary effluent filtered through biological activated carbon (O3/BAC) for 7-mo and tertiary effluent for 38-mo. Each column pair was operated in series, where the first column was operated for a shorter residence time and the second column for a longer residence time. Residence times tested were 5-d, 28-d, 30-d, 58-d, 60-d, 150-d and 180-d. For the last 38-mo, both pairs of columns had a residence time of 30-d in the first column and the total residence time of the two pairs was 150 and 180-d, respectively. Testing showed both of these pairs had the same long-term performance. The column pairs with a 150 to 180-d residence time, which were both fed tertiary effluent, reached an effluent total organic carbon (TOC) of 1.8 mg/L. Column pairs with a 28 to 30-d residence time, which were fed tertiary, ozonated, and O3/BAC effluent, reached effluent TOCs of 2.3, 2.1 and 1.8 mg/L respectively. In the latter, some TOC removal was shifted from the soil columns to the BAC. During the last 38 months of testing, using tertiary effluent as the source water, a series of sampling events was performed throughout the soil column system for N-nitrosodimethylamine (NDMA) and chemicals of emerging concern (CECs). NDMA was substantially reduced in all the columns, with a median value of 3 ng/L after 30-d and <2 ng/L after both 150 and 180-d. Twenty-one CECs were found in the majority of tertiary effluent samples, twelve of which were attenuated by the soil columns and the remaining were not. Chemicals found to be recalcitrant were 4-nonylphenol, acesulfame-k, carbamazepine, lidocaine, primidone, simazine, sucralose, sulfamethoxazole, and TCEP. Using excitation-emission matrix (EEM) techniques, soluble microbial products (SMP) peak characteristic of effluent organic matter (EfOM) is nearly eliminated after a 30-d hydraulic retention time (HRT) and completely eliminated in the 150/180-d samples. The intensity of the other peaks is significantly reduced as well, resulting in an EEM much like that of natural groundwater.

Late season pharmaceutical fate in wetland mesocosms with and without phosphorous addition.

The fate of six human-use drugs was assessed and predicted in mesocosms designed to mimic shallow constructed wetlands during the onset of fall and senescence. Mesocosms were monitored for 28 days after the addition of carbamazepine, clofibric acid, fluoxetine and naproxen (nominal initial concentrations of 5 µg/L each), sulfamethoxazole, and sulfapyridine (nominal initial concentrations of 150 µg/L each), with and without phosphorous (P) addition at 1.6 mg/L. We hypothesized that addition of P would stimulate primary productivity and enhance removal of pharmaceuticals from the water column. Carbamazepine, clofibric acid, fluoxetine, and naproxen had half-lives of 8.7, 11, 1.5, and 2.5, and 8.6, 11.0, 1.4, and 2.5 days in treatments with and without P amendment, respectively. Sulfamethoxazole and sulfapyridine had half-lives of 17 and 4.9 days in mesocosms with P amendment and 17 and 4.7 days without amendment. A concurrent pulse of P with pharmaceuticals did not significantly enhance the removal of these compounds. Predicted half-lives from modeling efforts were consistent with observed values, with photolysis the greatest contributor to chemical attenuation.

Carbamazepine derivatives with P2X4 receptor-blocking activity.

Antagonists for the P2 receptor subtype P2X4, an ATP-activated cation channel receptor, have potential as novel drugs for the treatment of neuropathic pain and other inflammatory diseases. In the present study, a series of 47 carbamazepine derivatives including 32 novel compounds were designed, synthesized, and evaluated as P2X4 receptor antagonists. Their potency to inhibit ATP-induced calcium influx in 1321N1 astrocytoma cells stably transfected with the human P2X4 receptor was determined. Additionally, species selectivity (human, rat, mouse) and receptor subtype selectivity (P2X4 vs P2X1, 2, 3, 7) were investigated for selected derivatives. The most potent compound of the present series, which exhibited an allosteric mechanism of P2X4 inhibition, was N,N-diisopropyl-5H-dibenz[b,f]azepine-5-carboxamide (34, IC50 of 3.44µM). The present study extends the so far very limited knowledge on structure-activity relationships of P2X4 receptor antagonists.

Ozonation products of carbamazepine and their removal from secondary effluents by soil aquifer treatment--indications from column experiments.

Ozonation is known as an efficient treatment to reduce the concentration of many trace organic compounds from WWTP effluents, but the formation of unknown and possibly persistent and toxic transformation products has to be considered. In this paper tertiary treatment of wastewater by the combination of ozone and soil aquifer treatment was investigated with respect to the removal of the antiepileptic drug carbamazepine (CBZ, 10 µg/L) and its transformation products. Batch tests and pilot experiments confirmed efficient removal of carbamazepine from secondary effluent by ozone. With typical ozone consumption of 0.7 mg O3/mg DOC0, approx. 50% of the transformed CBZ was detected as its primary product 1-(2-benzaldehyde)-4-hydro-(1H,3H)-quinazoline-2-one (BQM). Structure proposals and a formation pathway were elaborated for a total of 13 ozonation products of CBZ. In subsequent biological treatment BQM turned out to be more effectively biodegraded than CBZ. Its aldehyde group was quickly oxidized to a carboxylic acid (BaQM), which was removed in sand column experiments. Most of the minor ozonation products of CBZ persisted in sand column experiments with residence times of 5-6 days. Non-target screening of column effluent revealed no formation of persistent biotransformation products.

In situ monitoring of carbamazepine-nicotinamide cocrystal intrinsic dissolution behaviour.

Cocrystals have shown huge potential to improve the dissolution rate and absorption of a poorly water soluble drug. However, solution mediated phase transformation of cocrystals could greatly reduce the enhancement of its apparent solubility and dissolution rate. The aim of this study is to gain a deep understanding of the phase transition behaviour of cocrystals during dissolution and to investigate the improvement of dissolution rate. Dissolution and transformation behaviour of carbamazepine-nicotinamide (CBZ-NIC) cocrystal, physical mixture and different forms of carbamazepine: form I (CBZ I), form III (CBZ III) and dihydrate (CBZ DH) were studied by different in situ techniques of UV imaging and Raman spectroscopy. It has been found that compared with CBZ III and I, the rate of intrinsic dissolution rate (IDR) of CBZ-NIC cocrystal decreases slowly during dissolution, indicating the rate of crystallisation of CBZ DH from the solution is slow. In situ solid-state characterisation has shown the evolution of conversion of CBZ-NIC cocrystal and polymorphs to its dihydrate form. The study has shown that in situ UV imaging and Raman spectroscopy with a complementary technique of SEM can provide an in depth understanding during dissolution of cocrystals.

In vitro and in vivo evaluation of ordered mesoporous silica as a novel adsorbent in liquisolid formulation.

BACKGROUND: A liquisolid technique has been reported to be a new approach to improve the release of poorly water-soluble drugs for oral administration. However, an apparent limitation of this technique is the formulation of a high dose because a large amount of liquid vehicle is needed, which finally results in a low-dose liquisolid formulation. Silica as an absorbent has been used extensively in liquisolid formulations. Although nanoparticle silica can be prepared and used to improve liquid adsorption capacity, loading a high dose of drug into a liquisolid is still a challenge. With the aim of improving adsorption capacity and accordingly achieving high drug loading, ordered mesoporous silica with a high surface area and narrow pore size distribution was synthesized and used in a liquisolid formulation. METHODS: Ordered mesoporous silica was synthesized and its particle size and morphology were tailored by controlling the concentration of cetyltrimethyl ammonium bromide. The ordered mesoporous silica synthesized was characterized by transmission electron microscopy, scanning electron microscopy, Fourier transform infrared spectroscopy, small-angle x-ray diffraction, wide angle x-ray diffraction, and nitrogen adsorption-desorption measurements. The liquid adsorption capacity of ordered mesoporous silica was subsequently compared with that of conventional silica materials using PEG400 as the model liquid. Carbamazepine was chosen as a model drug to prepare the liquisolid formulation, with ordered mesoporous silica as the adsorbent material. The preparation was evaluated and compared with commercially available fast-release carbamazepine tablets in vitro and in vivo. RESULTS: Characterization of the ordered mesoporous silica synthesized in this study indicated a huge Brunauer-Emmett-Teller surface area (1030 m(2)/g), an ordered mesoporous structure with a pore size of 2.8 nm, and high adsorption capacity for liquid compared with conventional silica. Compared with fast-release commercial carbamazepine tablets, drug release from the liquisolid capsules was greatly improved, and the bioavailability of the liquisolid preparation was enhanced by 182.7%. CONCLUSION: Ordered mesoporous silica is a potentially attractive adsorbent which may lead to a new approach for development of liquisolid products.

Discriminatory dissolution method for quality control measurements of carbamazepine immediate release tablets based on in vitro--in vivo investigations.

The purpose of this study was to identify a discriminatory dissolution method able to predict the in vivo performance of tablet formulations designed for carbamazepine (CBZ). After evaluation of dissolution medium and rotation speed using a 25 central composite design and investigation of the in vivo release behaviors in beagle dogs, the dissolution method of CBZ 100 mg tablets was validated using a USP apparatus II, at a rotation speed of 75 rpm, and 900 ml deaerated water with 0.5% sodium lauryl sulfate (w/v) as the dissolution medium. Dissolution profiles were evaluated by the Weibull parameters and the modified fit factor, ƒ^(1,area). The in vitro-in vivo relationship of CBZ tablets was examined. Compared with the results from the USP and Chinese Pharmacopoeia monograph, the proposed system provides a superior discriminatory method. Since the dissolution method in pharmacopoeia for CBZ tablets is unable to distinguish between a good and a bad product, the method presented here can be used for the quality control testing of CBZ tablets.

Physico-mechanical and stability evaluation of carbamazepine cocrystal with nicotinamide.

The focus of this investigation was to prepare the cocrystal of carbamazepine (CBZ) using nicotinamide as a coformer and to compare its preformulation properties and stability profile with CBZ. The cocrystal was prepared by solution cooling crystallization, solvent evaporation, and melting and cryomilling methods. They were characterized for solubility, intrinsic dissolution rate, chemical identification by Fourier transform infrared spectroscopy, crystallinity by differential scanning calorimetry, powder X-ray diffraction, and morphology by scanning electron microscopy. Additionally, mechanical properties were evaluated by tensile strength and Heckel analysis of compacts. The cocrystal and CBZ were stored at 40°C/94% RH, 40°C/75% RH, 25°C/60% RH, and 60°C to determine their stability behavior. The cocrystals were fluffy, with a needle-shaped crystal, and were less dense than CBZ. The solubility profiles of the cocrystals were similar to CBZ, but its intrinsic dissolution rate was lower due to the high tensile strength of its compacts. Unlike CBZ, the cocrystals were resistant to hydrate transformation, as revealed by the stability studies. Plastic deformation started at a higher compression pressure in the cocrystals than CBZ, as indicated by the high yield pressure. In conclusion, the preformulation profile of the cocrystals was similar to CBZ, except that it had an advantageous resistance to hydrate transformation.

Fate and transport of carbamazepine in soil aquifer treatment (SAT) infiltration basin soils.

The transport and fate of the pharmaceutical carbamazepine (CBZ) were investigated in the Dan Region Reclamation Project (SHAFDAN), Tel-Aviv, Israel. Soil samples were taken from seven subsections of soil profiles (150 cm) in infiltration basins of a soil aquifer treatment (SAT) system. The transport characteristics were studied from the release dynamics of soil-resident CBZ and, subsequently, from applying a pulse input of wastewater containing CBZ. In addition, a monitoring study was performed to evaluate the fate of CBZ after the SAT. Results of this study indicate adsorption, and consequently retardation, in CBZ transport through the top soil layer (0-5 cm) and to a lesser extent in the second layer (5-25 cm), but not in deeper soil layers (25-150 cm). The soluble and adsorbed fractions of CBZ obtained from the two upper soil layers comprised 45% of the total CBZ content in the entire soil profile. This behavior correlated to the higher organic matter content observed in the upper soil layers (0-25 cm). It is therefore deduced that when accounting for the full flow path of CBZ through the vadose zone to the groundwater region, the overall transport of CBZ in the SAT system is essentially conservative. The monitoring study revealed that the average concentration of CBZ decreased from 1094 ± 166 ng L⁻¹ in the recharged wastewater to 560 ± 175 ng L⁻¹ after the SAT. This reduction is explained by dilution of the recharged wastewater with resident groundwater, which may occur as it flows to active reclamation wells.

Solvent-mediated solid phase transformations of carbamazepine: Effects of simulated intestinal fluid and fasted state simulated intestinal fluid.

Solvent-mediated transformations of carbamazepine (CBZ) anhydrate form III were investigated in Simulated Intestinal Fluid, a simple USP buffer medium, and in FaSSIF, which contains sodium taurocholate (STC) and lecithin, important surfactants that solubilize lipophilic drugs and lipids in the gastrointestinal tract. Raman spectroscopy (in situ) was utilized to reveal the connection between the changes in solid phase composition and dissolution rate while simultaneously detecting the solid state and the dissolved amount of CBZ. Initial dissolution rate was clearly higher in FaSSIF, while the solid phase data revealed that the crystallization of CBZ dihydrate was inhibited in both the dissolution media, albeit by different mechanisms. In SIF this inhibition was related to extensive needle growth, which impeded medium contact with the solid surface by forming a sterical barrier leading to retarded crystallization rates. Morphological changes from the needle-like dihydrate crystals to plate-like counterparts in FaSSIF, combined with the information that the transformation process was leveled off, evidenced strong hydrogen bonding behavior between the CBZ and STC molecules. These results underline the importance of biologically representative dissolution media in linking the in vitro dissolution results of solids that are capable of hydrate formation to their in vivo dissolution behavior.

The influence of various amphiphilic excipients on the physicochemical properties of carbamazepine-loaded microparticles.

The objective of the present study was to prepare multiple-unit formulations of carbamazepine (CBZ) using an emulsion congealing technique. CBZ-hydrogenated castor oil (HCO) (Cutina® HR) wax microparticles were prepared without organic solvents as an alternative to polymeric microparticles. The process involved emulsification and solidification of CBZ-HCO melt at a significantly low temperature (5°C). Five amphiphilic excipients (Pluronic F-68 (PL), Labrasol (LB), Gelucire 44/14 (GL 44/14), D-α-tocopheryl PEG 1000 succinate (TPGS) and Docusate sodium (DOSS) were added with the wax melt. The microparticles were characterized with respect to their particle size distribution, drug loading, morphological character, drug-excipient interaction, differential scanning calorimetry, Fourier-transform infra-red (FT-IR) and release properties. An average value for production yield was 83.45%. Evaluation of the release data indicates that the release mechanism from the prepared Cutina® HR microparticles follows both the Higuchi model of diffusion and anomalous release mechanism. Microparticles containing 5% Labrasol, TPGS and GL 44/14 had the highest extent of dissolution.

Can in vitro metabolism-dependent covalent binding data in liver microsomes distinguish hepatotoxic from nonhepatotoxic drugs? An analysis of 18 drugs with consideration of intrinsic clearance and daily dose.

In vitro covalent binding assessments of drugs have been useful in providing retrospective insights into the association between drug metabolism and a resulting toxicological response. On the basis of these studies, it has been advocated that in vitro covalent binding to liver microsomal proteins in the presence and the absence of NADPH be used routinely to screen drug candidates. However, the utility of this approach in predicting toxicities of drug candidates accurately remains an unanswered question. Importantly, the years of research that have been invested in understanding metabolic bioactivation and covalent binding and its potential role in toxicity have focused only on those compounds that demonstrate toxicity. Investigations have not frequently queried whether in vitro covalent binding could be observed with drugs with good safety records. Eighteen drugs (nine hepatotoxins and nine nonhepatotoxins in humans) were assessed for in vitro covalent binding in NADPH-supplemented human liver microsomes. Of the two sets of nine drugs, seven in each set were shown to undergo some degree of covalent binding. Among hepatotoxic drugs, acetaminophen, carbamazepine, diclofenac, indomethacin, nefazodone, sudoxicam, and tienilic acid demonstrated covalent binding, while benoxaprofen and felbamate did not. Of the nonhepatotoxic drugs evaluated, buspirone, diphenhydramine, meloxicam, paroxetine, propranolol, raloxifene, and simvastatin demonstrated covalent binding, while ibuprofen and theophylline did not. A quantitative comparison of covalent binding in vitro intrinsic clearance did not separate the two groups of compounds, and in fact, paroxetine, a nonhepatotoxin, showed the greatest amount of covalent binding in microsomes. Including factors such as the fraction of total metabolism comprised by covalent binding and the total daily dose of each drug improved the discrimination between hepatotoxic and nontoxic drugs based on in vitro covalent binding data; however, the approach still would falsely identify some agents as potentially hepatotoxic.

Rapid screening of glutathione-trapped reactive metabolites by linear ion trap mass spectrometry with isotope pattern-dependent scanning and postacquisition data mining.

The present study describes a novel integrated approach for rapid analysis of reactive metabolites with a linear ion trap mass spectrometer (LTQ). In this approach, an isotope pattern-dependent scanning method was applied to the data acquisition of glutathione (GSH)-trapped reactive metabolites. Recorded full-scan MS and MS/MS data sets were further processed with neutral loss filtering, product ion filtering, and extracted ion chromatographic analysis to search for protonated molecules and MS/MS spectra of GSH adducts. To evaluate the effectiveness and reliability of the approach, GSH adducts of carbamazepine, diclofenac, 4-ethylphenol, acetaminophen, p-cresol, and omeprazole were analyzed, which were formed in human liver microsome incubations fortified with a mixture of nonlabeled GSH and stable isotope-labeled GSH at a 1:0.8 ratio. Results demonstrate that the combination of the isotope pattern-dependent scanning with the postacquisition data mining was very effective in detecting low levels of GSH adducts, regardless of their fragmentation patterns. As compared to a neutral loss scanning method performed with a triple quadrupole mass spectrometer, the LTQ-based approach had several major advantages, including the superior selectivity and sensitivity in detecting different classes of GSH adducts and the higher throughput capability of the detection and MS/MS spectral acquisition of GSH adducts in a single LC/MS run. Overall, this analytical approach provides a simple and efficient means for screening for reactive metabolites using a linear ion trap LC/MS platform.