Fernanda de Lima Moreira

A non-michaelian behavior of the in vitro metabolism of the pentacyclic triterpene alfa and beta amyrins by employing rat liver microsomes

Pharmacological studies employing alpha and beta amyrin have demonstrated potential application in several biological activities suggesting their application as promising drugs. In the early drug development, metabolism studies may give important parameters regarding the efficacy and safety of the drug candidate. Therefore, the aim of this work was to determine the enzymatic kinetic parameters of these pentacyclic triterpenes. Chromatographic analyzes were performed using a Shimadzu GC-MS system. The resolution of amyrins was achieved with a DB5-MS column of 0.25 μM film thickness, 30.0 cm length and 0.25 mm diameter. At this condition, the retention times of beta- and alpha-amyrin were 21.3 and 20.2 min, respectively. The proposed method showed to be linear over the concentration range of 0.16-42.18 μM for beta amyrin and 0.11-28.12 μM for alpha amyrin. The lowest concentration quantified by the validated method was 0.16 μM for beta and 0.11 μM for alpha amyrin. The stability study showed that amyrins were stable at room temperature for 12h and at 37°C for 1h. The absolute recovery of the amyrin isomers from the rat microsome was 54.3-59.2%. The enzymatic kinetics presented sigmoidal plots. It was observed a Vmax=0.698 ± 0.022 μmol/mg protein/min, S50=4.4 μM and Hill coefficient of 2.7 ± 0.17 for alpha amyrin and a Vmax=0.775 ± 0.034 μmol/mg protein/min, S50=7.0 μM and Hill coefficient of 2.5 ± 0.21 for beta amyrin. The obtained results give the first clues regarding amyrin metabolism and suggests a more detailed study conducted employing isolated CYP isoforms.

In vitro metabolism of monensin A: microbial and human liver microsomes models

Abstract 1. Monensin A, an important antibiotic ionophore that is primarily employed to treat coccidiosis, selectively complexes and transports sodium cations across lipid membranes and displays a variety of biological properties. 2. In this study, we evaluated the fungi Cunninghamella echinulata var. elegans ATCC 8688A, Cunninghamella elegans NRRL 1393 ATCC 10028B and human hepatic microsomes as CYP-P450 models to investigate the in vitro metabolism of monensin A and compare the products with the metabolites produced in vivo. 3. Mass spectrometry analysis of the products from these model systems revealed the formation of three metabolites: 3-O-demethyl monensin A, 12-hydroxy monensin A and 12-hydroxy-3-O-demethyl monensin A. We identified these products by tandem mass spectrometry and through comparison with the in vivo metabolites. 4. This analysis demonstrated that the model systems produce the same metabolites found in in vivo studies, thus they could be used to predict the metabolism of monensin A. Furthermore, we verified that liquid chromatography coupled to mass spectrometry is a powerful tool to study the in vitro metabolism of drugs, because it allows the successful identifications of several derivatives from different metabolic models.

Metabolic profile and safety of piperlongumine

Piperlongumine (PPL), a natural plant product, has been extensively studied in cancer treatment going up on clinical trials. Since the first report related to its use on cancer research (in 2011) around 80 papers have been published in less than 10 years, but a gap still remaining. There are no metabolism studies of PPL in human organism. For the lack of a better view, here, the CYP450 in vitro oxidation of PPL was described for the first time. In addition, the enzymatic kinetic data, the predicted in vivo parameters, the produced metabolites, the phenotyping study and possible piperlongumine-drug interactions in vivo is presented.

In vitro enantioselective human liver microsomal metabolism and prediction of in vivo pharmacokinetic parameters of tetrabenazine by DLLME-CE☆

A new capillary electrophoresis method for the enantioselective analysis of cis- and trans- dihydrotetrabenazine (diHTBZ) after in vitro metabolism by human liver microsomes (HLMs) was developed. The chiral electrophoretic separations were performed by using tris-phosphate buffer (pH 2.5) containing 1% (w/v) carboxymethyl-β-CD as background electrolyte with an applied voltage of +15kV and capillary temperature kept at 15°C. Dispersive liquid-liquid microextraction was employed to extract the analytes from HLMs. Dichloromethane was used as extraction solvent (75μL) and acetone as disperser solvent (150μL). The method was validated according to official guidelines and showed to be linear over the concentration range of 0.29-19.57μmolL(-1) (r=0.9955) for each metabolite enantiomer. Within- and between-day precision and accuracy evaluated by relative standard deviation and relative error were lower than 15% for all enantiomers. The stability assay showed that the analytes kept stable under handling, storage and in metabolism conditions. After method validation, an enantioselective in vitro metabolism and in vivo pharmacokinetic prediction was carried out. This study showed a stereoselective metabolism and the observed kinetic profile indicated a substrate inhibition behavior. DiHTBZ enantiomers were catalyzed mainly by CYP2C19 and the predicted clearance suggests that liver metabolism is the main route for TBZ elimination which supports the literature data.

Analytical Methods for the Determination of Rosuvastatin in Pharmaceutical Formulations and Biological Fluids: A Critical Review

ABSTRACT Rosuvastatin calcium (ROS), (Figure 1) belongs to the “statins” group, which is the 3-hydroxy-3-methylglutaryl coenzyme A (HMG-CoA) reductase inhibitor. This drug is indicated for dyslipidemias treatment and can help to decrease the level of “bad cholesterol” and can consequently reduce the development of atherosclerosis and the risk of heart diseases. ROS was developed by Astra-Zeneca and it was approved in 2003 by the FDA in the United States. In 2015, under the trade name Crestor®, it was the fourth largest selling drug in the United States with sales above

In vitro metabolism studies of natural products are crucial in the early development of medicines

5 billion. This study presents a literature review of analytical methods for the quantification of ROS in pharmaceutical preparations and biological fluids. The major analytical methods described in this study for ROS were spectrophotometry, high-performance liquid chromatography (HPLC) coupled to ultraviolet (UV) detection, and tandem mass spectrometry (LC–MS/MS).

Lopinavir/ritonavir treatment increases the placental transfer of bupivacaine enantiomers in human immunodeficiency virus‐infected pregnant women

Finding new drugs is imperative especially in the case of diseases such as serious infections and cancer, for which a gap in drug development exists. Plant-derived compounds display large structural diversity and constitute a source of novel medicinal agents. However, some drug candidates have failed in advanced stages of clinical and commercial use. In vitro tools have been effectively used to anticipate and to explore undesirable drug properties during early drug development. In this review, we address the purpose and advantages of conducting metabolic studies on natural products, and we describe how in vitro metabolism methods have contributed to delineating plant-based drug candidates. We also deal with the applications of in vitro metabolism studies on natural product-based drug development, which include characterization of the metabolites, investigation into enzymatic kinetics, prediction of drug clearance, phenotyping studies, and evaluation of natural product–drug interactions.

In vitro metabolism of the lignan (-)-grandisin, an anticancer drug candidate, by human liver microsomes.

The present study evaluated the placental transfer and amniotic fluid distribution of bupivacaine enantiomers in health pregnant women and in human immunodeficiency virus (HIV)‐infected pregnant women receiving epidural anaesthesia for caesarean section.