Investigating Bacopa monnieri L. Therapeutic Potential for the Treatment of Neurological Diseases.

The popular perennial creeping plant known as Bacopa monnieri (also known as Brahmi) is being utilized in the Indian Ayurvedic medicine practice. It has a variety of bioactive phytoconstituents that have been used therapeutically to treat a number of serious illnesses. Ancient Vedic scholars used this herb because of its pharmacological effects, particularly as a nerve booster and nootropic supporter. However, it is vital to comprehend the active phytochemical components of Bacopa monnieri extract (BME) and their molecular mechanisms in order to better grasp the effect of BME on neurological illnesses and diseases. Understanding its active phytochemical constituents and their molecular processes is essential. Numerous clinical investigations indicated that BME may have neuroprotective benefits, so it is worthwhile to re-evaluate this wellknown plant. Here, we focused on neurological problems as we examined the pharmacological and phytochemical characteristics of BME. For their effective usage in neuroprotection and cognition, many clinical concerns and the synergistic potential of Bacopa extract have been investigated. Alzheimer's disease is a neurological condition caused by the production of reactive oxygen species, which also causes amyloid-beta (A) and tau protein aggregation and increases neuro-inflammation and neurotoxicity. Our review offers a more indepth molecular understanding of the neuroprotective functions of BME, which can also be connected to its therapeutic management of neurological illnesses and cognitive-improving effects.

Targeted Protein Degraders- The Druggability Perspective.

Targeted Protein degraders (TPDs) show promise in harnessing cellular machinery to eliminate disease-causing proteins, even those previously considered undruggable. Especially if protein turnover is low, targeted protein removal bestows lasting therapeutic effect over typical inhibition. The demonstrated safety and efficacy profile of clinical candidates has fueled the surge in the number of potential candidates across different therapeutic areas. As TPDs often do not comply with Lipinski's rule of five, developing novel TPDs and unlocking their full potential requires overcoming solubility, permeability and oral bioavailability challenges. Tailored in-vitro assays are key to precise profiling and optimization, propelling breakthroughs in targeted protein degradation.

Friedelin: Structure, Biosynthesis, Extraction, and Its Potential Health Impact.

Pharmaceutical companies are investigating more source matrices for natural bioactive chemicals. Friedelin (friedelan-3-one) is a pentacyclic triterpene isolated from various plant species from different families as well as mosses and lichen. The fundamental compounds of these friedelane triterpenoids are abundantly found in cork tissues and leaf materials of diverse plant genera such as Celastraceae, Asteraceae, Fabaceae, and Myrtaceae. They possess many pharmacological effects, including anti-inflammatory, antioxidant, anticancer, and antimicrobial activities. Friedelin also has an anti-insect effect and the ability to alter the soil microbial ecology, making it vital to agriculture. Ultrasound, microwave, supercritical fluid, ionic liquid, and acid hydrolysis extract friedelin with reduced environmental impact. Recently, the high demand for friedelin has led to the development of CRISPR/Cas9 technology and gene overexpression plasmids to produce friedelin using genetically engineered yeast. Friedelin with low cytotoxicity to normal cells can be the best phytochemical for the drug of choice. The review summarizes the structural interpretation, biosynthesis, physicochemical properties, quantification, and various forms of pharmacological significance.

Anti-inflammatory and Anti-arthritic Properties of Mucuna gigantea Plant Extracts: Establishing by Molecular Docking Study.

Background Mucuna giganteais a traditional plant reported in the management of nervous disorders, male infertility, etc., and also exhibits aphrodisiac, anti-oxidant, and anti-diabetic properties. Very few studies are conducted on Mucuna gigantea. It has not been pharmacologically evaluated for rheumatoid arthritis (RA). In RA, the body's natural defence mechanism gets confused and begins to target the healthy tissues in the body, which leads to joint pain, swelling, bone erosion, and joint stiffness. It is a condition that is classified as an auto-immune disorder. Methods In-silico docking depicted that beta-sitosterol is present in Mucuna gigantea out of ligand library prepared based on a literature survey using computational analysis. Inflammation was induced by carrageen and chronic inflammation was induced by Freund's complete adjuvant in the plantar surface of the rats. The petroleum ether, ethanolic and aqueous extracts in three divided doses (75, 150, and 300 mg/kg) were administered orally. Diclofenac sodium (10 mg/kg), prednisolone (5 mg/kg), and methotrexate (0.5 mg/kg) were used as standard. The statistical significance between means was analyzed using one-way ANOVA, followed by Dunnett's multiple range test. The values are expressed as mean ± SD for each group (n=6), and aP<0.0001, bP<0.001, and cP<0.05 were compared with a negative control group. Results Ethanolic and petroleum ether extracts showed a statistically significant aP<0.0001 effect at 3hr with 300mg/kg effect in analgesic activity, whereas aqueous extracts showed statistically significant aP<0.0001 effect at 1.5hr with 150 and 300mg/kg. In the carrageen-induced model, all three extracts at 300 mg/kg showed a statistically significant aP<0.0001 effect from 2- 4hr. In Freund's adjuvant model, all three extracts at all doses showed a statistically significant aP<0.0001 effect. Also, Mucuna gigantea remarkably ameliorated altered WBCs, rheumatoid factor, and positively modified radiographic and histopathological changes. Conclusion Taken together, these results support the traditional use of Mucuna gigantea as a potent anti-inflammatory and anti-arthritic agent that may be proposed for rheumatoid arthritis treatment.

Advances in Animal Models and Cutting-Edge Research in Alternatives: Proceedings of the Third International Conference on 3Rs Research and Progress, Vishakhapatnam, 2022.

Animal experimentation has been integral to drug discovery and development and safety assessment for many years, since it provides insights into the mechanisms of drug efficacy and toxicity (e.g. pharmacology, pharmacokinetics and pharmacodynamics). However, due to species differences in physiology, metabolism and sensitivity to drugs, the animal models can often fail to replicate the effects of drugs and chemicals in human patients, workers and consumers. Researchers across the globe are increasingly applying the Three Rs principles by employing innovative methods in research and testing. The Three Rs concept focuses on: the replacement of animal models (e.g. with in vitro and in silico models or human studies), on the reduction of the number of animals required to achieve research objectives, and on the refinement of existing experimental practices (e.g. eliminating distress and enhancing animal wellbeing). For the last two years, Oncoseek Bio-Acasta Health, a 3-D cell culture-based cutting-edge translational biotechnology company, has organised an annual International Conference on 3Rs Research and Progress. This series of global conferences aims to bring together researchers with diverse expertise and interests, and provides a platform where they can share and discuss their research to promote practices according to the Three Rs principles. In November 2022, the 3rd international conference, Advances in Animal Models and Cutting-Edge Research in Alternatives, took place at the GITAM University in Vishakhapatnam (AP, India) in a hybrid format (i.e. online and in-person). These conference proceedings provide details of the presentations, which were categorised under five different topic sessions. It also describes a special interactive session on in silico strategies for preclinical research in oncology, which was held at the end of the first day.

The Anti-inflammatory and Anti-arthritis Activity of Garcinia travancorica in a Rat Model: A Proof of Concept with Computational Studies.

Background Rheumatoid arthritis is a progressive disease of human joints characterized by severe pain, stiffness, and tissue damage at the local site. Bone and cartilaginous tissue damage at the synovial joints is initiated by the production of autoantibody induced by inflammatory signaling through cytokines. Objective This study aimed to evaluate the efficacy of Garcinia travancorica against acute and chronic inflammation in a rat model after designing the ligand library and target identification using computational analysis. Methods Acute inflammation was induced by carrageen, and chronic inflammation was induced by Freund's complete adjuvant in the plantar surface of the rats. The petroleum ether, ethanolic, and aqueous extracts in three divided doses (75 mg/kg, 150 mg/kg, and 300 mg/kg) were administered orally. Diclofenac sodium (10 mg/kg), prednisolone (5 mg/kg), and methotrexate (0.5 mg/kg) were used as standard. Various parameters were evaluated, and statistical significance between means was analyzed using one-way ANOVA, followed by Dunnett's multiple range test. Results: Docking-based in-silico screening of the ligand library has revealed the potential of Polyanxanthone-C as an anti-rheumatoid agent, which is supposed to deliver its therapeutic effect by synergistic targeting of interleukin-1, interleukin-6, and tumor necrosis factor receptor type-1. Conclusion: This plant has the potential to be used in the treatment of arthritis-related disorders.

Cytochrome P450 and P-gp Mediated Herb-Drug Interactions and Molecular Docking Studies of Garcinol.

Garcinol is an active constituent of Garcinia indica and Garcinia cambogia. Recent studies have proven that garcinol has anti-inflammatory, anti-cancer, and anti-oxidant activities. The objective of this study was to evaluate the inhibitory effects of garcinol on the activities of the drug metabolizing cytochrome P450 (CYP) isozymes to predict potential herb-drug interactions with co-administered drugs. Garcinol was incubated with a mixture of rat liver microsomes and eight CYP probe substrate cocktail under optimized incubation conditions and the samples were analyzed using a validated method on LC-MS/MS. Garcinol showed strong inhibition with IC50 values of CYP1A2 (7.6 µM), CYP2C9 (8.0 µM), CYP2B6 (2.1 µM), CYP2D6 (9.5 µM), and CYP3A4 (5.1 µM), respectively, and moderate inhibition towards CYP2C19 (16.4 µM) and CYP2E1 (19.0 µM). Molecular docking studies were performed on garcinol against the active sites of CYP2B6 and CYP3A4 proteins. These results further confirmed that the inhibitory activity of garcinol occurred by occupying the active sites of these human CYPs and by making favorable interactions with its key residues. In-vivo CYP inhibition studies were carried out in Sprague-Dawley rats. These results suggest garcinol may cause herb-drug interactions, mediated by inhibition of CYPs involved in drug metabolism in-vivo by altering the pharmacokinetic parameters like AUC and Cmax in a clinically significant manner. Garcinol was found to upregulate the expression and activity of P-gp in western blotting study and P-gp inhibition study in-vivo. These findings give a clear understanding to predict potential herb-drug/drug-drug interactions of garcinol for safe clinical use in future.

Three-dimensional aspects of formulation excipients in drug discovery: a critical assessment on orphan excipients, matrix effects and drug interactions.

Formulation/pharmaceutical excipients play a major role in formulating drug candidates, with the objectives of ease of administration, targeted delivery and complete availability. Many excipients used in pharmaceutical formulations are orphanized in preclinical drug discovery. These orphan excipients could enhance formulatability of highly lipophilic compounds. Additionally, they are safe in preclinical species when used below the LD50 values. However, when the excipients are used in formulating compounds with diverse physico-chemical properties, they pose challenges by modulating study results through their bioanalytical matrix effects. Excipients invariably present in study samples and not in the calibration curve standards cause over-/under- estimation of exposures. Thus, the mechanism by which excipients cause matrix effects and strategies to nullify these effects needs to be revisited. Furthermore, formulation excipients cause drug interactions by moderating the pathways of drug metabolizing enzymes and drug transport proteins. Although it is not possible to get rid of excipient driven interactions, it is always advised to be aware of these interactions and apply the knowledge to draw meaningful conclusions from study results. In this review, we will comprehensively discuss a) orphan excipients that have wider applications in preclinical formulations, b) bioanalytical matrix effects and possible approaches to mitigating these effects, and c) excipient driven drug interactions and strategies to alleviate the impacts of drug interactions.

Three-dimensional aspects of formulation excipients in drug discovery:a critical assessment on orphan excipients, matrix effects and drug interactions / 药物分析学报

Formulation/pharmaceutical excipients play a major role in formulating drug candidates, with the ob-jectives of ease of administration, targeted delivery and complete availability. Many excipients used in pharmaceutical formulations are orphanized in preclinical drug discovery. These orphan excipients could enhance formulatability of highly lipophilic compounds. Additionally, they are safe in preclinical species when used below the LD50 values. However, when the excipients are used in formulating compounds with diverse physico-chemical properties, they pose challenges by modulating study results through their bioanalytical matrix effects. Excipients invariably present in study samples and not in the cali-bration curve standards cause over-/under- estimation of exposures. Thus, the mechanism by which excipients cause matrix effects and strategies to nullify these effects needs to be revisited. Furthermore, formulation excipients cause drug interactions by moderating the pathways of drug metabolizing en-zymes and drug transport proteins. Although it is not possible to get rid of excipient driven interactions, it is always advised to be aware of these interactions and apply the knowledge to draw meaningful conclusions from study results. In this review, we will comprehensively discuss a) orphan excipients that have wider applications in preclinical formulations, b) bioanalytical matrix effects and possible ap-proaches to mitigating these effects, and c) excipient driven drug interactions and strategies to alleviate the impacts of drug interactions.

Discovery and Characterization of Potent Pan-Genotypic HCV NS5A Inhibitors Containing Novel Tricyclic Central Core Leading to Clinical Candidate.

The identification of a novel class of potent pan-genotypic NS5A inhibitors with good pharmacokinetic profile suitable for potential use in treating HCV infections is disclosed here. The present series of compounds are with less complex tricyclic central core, identified through a systematic SAR study carried out on biphenyl moiety. The SAR outcome has confirmed the requirement of near planar and linear conformation of the molecule to achieve the best pan-genotypic activity. In addition, SAR with substituted imidazoles on improvement of antiviral activity is disclosed. The newly identified compounds 12, 16, 19-21 have shown desirable pharmacokinetic profiles with a favorable uptake of compounds in liver and maintained a significant concentration for up to 8 h in the liver. In addition, compounds 20 and 21 have shown superior pan-genotypic anti-HCV activity compared to ledipasvir and daclatasvir. Additional characterization and preliminary safety assessment resulted in the identification of compound 20 as a potential clinical candidate.

Galeterone and The Next Generation Galeterone Analogs, VNPP414 and VNPP433-3ß Exert Potent Therapeutic Effects in Castration-/Drug-Resistant Prostate Cancer Preclinical Models In Vitro and In Vivo.

These studies compared the efficacies of our clinical agent galeterone (Gal) and the FDA-approved prostate cancer drug, enzalutamide (ENZ) with two lead next generation galeterone analogs (NGGAs), VNPP414 and VNPP433-3ß, using prostate cancer (PC) in vitro and in vivo models. Antitumor activities of orally administered agents were also assessed in CWR22Rv1 tumor-bearing mice. We demonstrated that Gal and NGGAs degraded AR/AR-V7 and Mnk1/2; blocked cell cycle progression and proliferation of human PC cells; induced apoptosis; inhibited cell migration, invasion, and putative stem cell markers; and reversed the expression of epithelial-to-mesenchymal transition (EMT). In addition, Gal/NGGAs (alone or in combination) also inhibited the growth of ENZ-, docetaxel-, and mitoxantrone-resistant human PC cell lines. The NGGAs exhibited improved pharmacokinetic profiles over Gal in mice. Importantly, in vivo testing showed that VNPP433-3ß (at 7.53-fold lower equimolar dose than Gal) markedly suppressed (84% vs. Gal, 47%; p < 0.01) the growth of castration-resistant PC (CRPC) CWR22Rv1 xenograft tumors, with no apparent host toxicity. ENZ was ineffective in this CRPC xenograft model. In summary, our findings show that targeting AR/AR-V7 and Mnk1/2 for degradation represents an effective therapeutic strategy for PC/CRPC treatment and supports further development of VNPP433-3ß towards clinical investigation.

Investigation of the inhibition of eight major human cytochrome P450 isozymes by a probe substrate cocktail in vitro with emphasis on CYP2E1.

1. A protocol has been developed and validated for the high-throughput screening of eight major human cytochrome P450 (CYP) isozymes inhibition (CYP 1A2, 2C9, 2C19, 2D6, 3A4, 2B6, 2C8 and 2E1) using an in vitro probe cocktail containing eight substrates by overcoming the unfavorable effect of assay conditions on CYP2E1 inhibition data. 2. The cocktail consisting of selective probe substrates like tacrine (CYP1A2), diclofenac (CYP2C9), S-mephenytoin (CYP2C19), dextromethorphan (CYP2D6), midazolam (CYP3A4), bupropion (CYP2B6), paclitaxel (CYP2C8) and chlorzoxazone (CYP2E1) was incubated with human liver microsomes. 3. The method was investigated by incubating well-known CYP inhibitors {alphanaphthoflavone (CYP1A2), sulfaphenazole (CYP2C9), N-3-benzylnirvanol (CYP2C19), quinidine (CYP2D6), ketoconazole (CYP3A4), ticlopidine (CYP2B6), quercetin (CYP2C8) and 4-methylpyrazole (CYP2E1)} with the substrate cocktail. A fast gradient liquid chromatography tandem mass spectrometry (LC-MS/MS) was used for this study. 4. The IC50 values determined for typical CYP inhibitors were reproducible and consistent with those in the literature. DMSO has significant effect and itself inhibits CYP2E1. DMSO should not exceed 0.1% for the determination of reliable CYP2E1 inhibition profile. This cocktail assay offers an efficient and robust method to determine the CYP450 isoforms inhibition profiles of large numbers of compounds in a quick turnaround time.

Design and synthesis of functionalized piperazin-1yl-(E)-stilbenes as inhibitors of 17α-hydroxylase-C17,20-lyase (Cyp17).

The synthesis of steroid hormones is critical to human physiology and improper regulation of either the synthesis of these key molecules or activation of the associated receptors can lead to disease states. This has led to intense interest in developing compounds capable of modulating the synthesis of steroid hormones. Compounds capable of inhibiting Cyp19 (Aromatase), a key enzyme in the synthesis of estrogens, have been successfully employed as breast cancer therapies, while inhibitors of Cyp17 (17α-hydroxylase-17,20-lyase), a key enzyme in the synthesis of glucocorticoids, mineralocorticoids and steroidal sex hormones, are a key component of prostate cancer therapy. Inhibition of CYP17 has also been suggested as a possible target for the treatment of Cushing Syndrome and Metabolic Syndrome. We have identified two novel series of stilbene based CYP17 inhibitors and demonstrated that exemplary compounds in these series have pharmacokinetic properties consistent with orally delivered drugs. These findings suggest that compounds in these classes may be useful for the treatment of diseases and conditions associated with improper regulation of glucocorticoids synthesis and glucocorticoids receptor activation.

Design, synthesis, and evaluation of (2S,4R)-Ketoconazole sulfonamide analogs as potential treatments for Metabolic Syndrome.

Metabolic Syndrome, also referred to as 'Syndrome X' or 'Insulin Resistance Syndrome,' remains a major, unmet medical need despite over 30years of intense effort. Recent research suggests that there may be a causal link between this condition and abnormal glucocorticoid processing. Specifically, dysregulation of the hypothalamic-pituitary-adrenocortical (HPA) axis leads to increased systemic cortisol concentrations. Cushing' syndrome, a disorder that is also typified by a marked elevation in levels of cortisol, produces clinical symptomology that is similar to those observed in MetS, and they can be alleviated by decreasing circulating cortisol concentrations. As a result, it has been suggested that decreasing systemic cortisol concentration might have a positive impact on the progression of MetS. This could be accomplished through inhibition of enzymes in the cortisol synthetic pathway, 11ß-hydroxylase (Cyp11B1), 17α-hydroxylase-C17,20-lyase (Cyp17), and 21-hydroxylase (Cyp21). We have identified a series of novel sulfonamide analogs of (2S,4R)-Ketoconazole that are potent inhibitors of these enzymes. In addition, selected members of this class of compounds have pharmacokinetic properties consistent with orally delivered drugs, making them well suited to further investigation as potential therapies for MetS.

Breaking the sensitivity limitations of cytochrome P450 oxidation product: dansyl chloride derivatisation of 4-OH mephenytoin, a CYP2C19 metabolite and its application to in vitro CYP inhibition assay.

A rapid selective and sensitive liquid chromatography/tandem mass spectrometry (LC-MS/MS) method was developed for the quantitative determination of derivatised cytochrome P450-2C19 oxidation product (dansyl-4-OH mephenytoin) and its underivatised form (4-OH mephenytoin). Samples were anaysed on C18 column (Waters Xbridge, 50 mm×4.6 mm, 3.5 µm particle size) with the mobile phase consisting of 0.1% formic acid in water and 0.1% formic acid in acetonitrile. A gradient method with a short run time of 2.5 min and 3.5 min was developed for the analysis of dansyl-4-OH mephenytoin and 4-OH mephenytoin, respectively. The standard curve was linear (r(2)=0.9972 for 4-OH mephenytoin; r(2)=0.9946 for dansyl-4-OH mephenytoin) over the concentration range of 0.16 to 40 ng/mL for both derivatised and underivatised forms. The CV (%) and relative error (RE) for inter and intraassay at three QC levels for dansyl-4-OH mephenytoin was 0.97-5.85% and -9.80 to 2.51%, respectively. Whereas, for 4-OH mephenytoin the CV (%) and RE (%) at three QC levels was 0.82-3.47% and -6.69 to -0.01%, respectively. The developed method was validated for various parameters such as linearity, precision & accuracy, extraction recovery, matrix effect, autosampler stability and was proved to be consistent across three QC levels with overall CV (%) less than 15. Dansylation helped in increasing the sensitivity of hydroxy mephenytoin by 100-200 fold. Given the simplicity involved in derivatisation process, we believe that this novel methodology will change the current approaches used for the enhancing the detection sensitivity of 4-OH mephenytoin.

Periosteal Pedicle Flap Harvested during Vestibular Extension for Root Coverage.

Root exposure along with inadequate vestibular depth is a common clinical finding. Treatment option includes many techniques to treat such defects for obtaining predictable root coverage. Normally, the vestibular depth is increased first followed by a second surgery for root coverage. The present case report describes a single-stage technique for vestibular extension and root coverage in a single tooth by using the Periosteal Pedicle Flap (PPF). This technique involves no donor site morbidity and allows for reflection of sufficient amount of periosteal flap tissue with its own blood supply at the surgical site, thus increasing the chances of success of root coverage with simultaneous increase in vestibular depth.

Liquid chromatography/tandem mass spectrometry method for quantitative estimation of solutol HS15 and its applications.

A rapid, sensitive and selective pseudoMRM (pMRM)-based method for the determination of solutol HS15 (SHS15) in rat plasma was developed using liquid chromatography/tandem mass spectrometry (LC-MS/MS). The most abundant ions corresponding to SHS15 free polyethyleneglycol (PEG) oligomers at m/z 481, 525, 569, 613, 657, 701, 745, 789, 833, 877, 921 and 965 were selected for pMRM in electrospray mode of ionization. Purity of the lipophilic and hydrophilic components of SHS15 was estimated using evaporative light scattering detector (ELSD). Plasma concentrations of SHS15 were measured after oral administration at 2.50 g/kg dose and intravenous administration at 1.00 g/kg dose in male Sprague Dawley rats. SHS15 has poor oral bioavailability of 13.74% in rats. Differences in pharmacokinetics of oligomers were studied. A novel proposal was conveyed to the scientific community, where formulation excipient could be analyzed as a qualifier in the analysis of new chemical entities (NCEs) to address the spiky plasma concentration profiles.

Liquid chromatography/tandem mass spectrometry method for quantitative estimation of solutol HS15 and its applications / 药物分析学报

A rapid, sensitive and selective pseudoMRM (pMRM)-based method for the determination of solutol HS15 (SHS15) in rat plasma was developed using liquid chromatography/tandem mass spectro-metry (LC–MS/MS). The most abundant ions corresponding to SHS15 free polyethyleneglycol (PEG) oligomers at m/z 481, 525, 569, 613, 657, 701, 745, 789, 833, 877, 921 and 965 were selected for pMRM in electrospray mode of ionization. Purity of the lipophilic and hydrophilic components of SHS15 was estimated using evaporative light scattering detector (ELSD). Plasma concentrations of SHS15 were measured after oral administration at 2.50 g/kg dose and intravenous administration at 1.00 g/kg dose in male Sprague Dawley rats. SHS15 has poor oral bioavailability of 13.74% in rats. Differences in pharmacokinetics of oligomers were studied. A novel proposal was conveyed to the scientific community, where formulation excipient could be analyzed as a qualifier in the analysis of new chemical entities (NCEs) to address the spiky plasma concentration profiles.

Pharmacokinetics and Excretion Studies on CDRI-85/92, an Antiulcer Proton Pump Inhibitor.

CDRI 85/92 is an antiulcer pharmacophore and a proton pump inhibitor, which is in an advanced stage of preclinical trials. In view of its importance, pharmacokinetic and excretion were studied in Sprague Dawley rats after administering 20 mg/kg oral and intravenous doses. The compound was detectable in the serum samples as early as 5 min post-oral administration. The compound was eliminated slowly from serum with an elimination half-life of 2.1 h. Following the 20 mg/kg oral dose, maximum serum concentration (C(max)) was found to be 469.28 ± 45.52 ng/ml after 1.0 h. Based on AUC values, the absolute bioavailability of the CDRI 85/92 was 70.5% after oral administration. It was found to be excreted in urine (~15% of the dose) in intravenously treated (bile duct cannulated as well as noncannulated) rats, whereas bile and feces depicted insignificant levels of the compound.