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IUCrData Apr 2023The title compound, CHNO, crystallizes with a disordered nitro group in twinned crystals. Both the meth-oxy group and the acetamide groups are nearly coplanar with the...
The title compound, CHNO, crystallizes with a disordered nitro group in twinned crystals. Both the meth-oxy group and the acetamide groups are nearly coplanar with the phenyl ring, and the C-N-C-O torsion angle [0.2 (4)°] is also insignificantly different from zero. Overall, the 12-atom meth-oxy-phenyl-acetamide group is nearly planar, with an r.m.s. deviation of 0.042 Å. The nitro group is twisted out of this plane by about 30°, disordered into two orientations with opposite senses of twist. In the crystal, the N-H group donates a hydrogen bond to a nitro oxygen atom, generating chains propagating in the [101] direction. The amide carbonyl oxygen atom is not involved in the hydrogen bonding.
PubMed: 37151207
DOI: 10.1107/S2414314623002985 -
European Journal of Clinical... Jun 2023Pregnancy-mediated physiological and biochemical changes contribute to alterations in the pharmacokinetics of certain drugs. There is a paucity of data on the systematic...
A cocktail probe approach to evaluate the effect of hormones on the expression and activity of CYP enzymes in human hepatocytes with conditions simulating late stage of pregnancy.
PURPOSE
Pregnancy-mediated physiological and biochemical changes contribute to alterations in the pharmacokinetics of certain drugs. There is a paucity of data on the systematic evaluation of the underlying mechanisms. The objective of the current study was to examine the impact of changes in circulating and tissue hormonal concentration during the late stage of pregnancy on the activity and expression of hepatic cytochrome P450 (CYP) enzymes using a cocktail probe approach.
METHODS
Freshly isolated primary human hepatocytes were incubated with third trimester physiologic (plasma) and projected liver (ten-fold higher) concentrations of female hormones: progesterone (2 µM), estradiol (0.3 µM), estriol (0.8 µM), estrone (0.2 µM), 17α-hydroxyprogesterone (0.1 µM), and human growth hormone (0.005 µM). The metabolic activity of the hepatocytes was assessed using a cocktail of isozyme-specific P450 probe substrates (CYP1A2 (phenacetin), CYP2C9 (diclofenac), CYP2C19 (S-mephenytoin), CYP2D6 (dextromethorphan), and CYP3A4 (testosterone)). A validated LC-MS/MS assay was used to measure the corresponding metabolite concentrations. CYP450 protein and mRNA levels were measured using western blot and qRT-PCR, respectively.
RESULTS
Female hormones at projected third-semester hepatic concentrations significantly enhanced mRNA and protein expression and increased the metabolic activity of CYP3A4. The expression and activity of other CYP450 enzymes studied were not affected by mixtures of female hormones at concentrations used.
CONCLUSION
The increased activity of CYP3A4 is consistent with the clinically observed increase in clearance of CYP3A4 substrates during pregnancy. Overall expression and activity of CYP450 isozymes are differentially regulated during pregnancy.
Topics: Humans; Female; Pregnancy; Cytochrome P-450 CYP3A; Chromatography, Liquid; Tandem Mass Spectrometry; Cytochrome P-450 Enzyme System; Hepatocytes; Hormones; Microsomes, Liver
PubMed: 37060457
DOI: 10.1007/s00228-023-03489-1 -
Frontiers in Pediatrics 2023drug exposure is a significant public health threat to the well-being and normal development of the neonate. Recently, testing of umbilical cord tissue (UCT) has been...
drug exposure is a significant public health threat to the well-being and normal development of the neonate. Recently, testing of umbilical cord tissue (UCT) has been employed to measure illicit drug exposure, as drugs used by the mother during the third trimester may be retained in the UCT. Focus has also been given to potential adverse health effects among drug users, resulting from exposure to pharmacologically active adulterants and cutting agents in the street drug supply. The effects of these substances have not been well studied in humans, nor has their presence been demonstrated as a means for assessing adverse health effects in the neonate. Here, we describe the application of a novel test method to analyze UCT for the presence of more than 20 common adulterating/cutting substances via LC/Q-TOF. In total, 300 de-identified UCT samples were analyzed-all had previously tested positive for cocaine or opiates. Generally, the positivity rates of individual compounds were similar between the Cocaine and Opiates Subgroups, apart from levamisole, xylazine, dipyrone (metabolites), and promethazine. Many of the adulterants used in the street drug supply do have legitimate medicinal/therapeutic uses, including several of the compounds most frequently detected in this study. Caffeine and lidocaine were the most frequently identified compounds both individually (>70% each) and in combination with each other. Alternatively, levamisole, an adulterant with no legitimate therapeutic use, was present in 12% of cases. Importantly, this data demonstrates that the detection of traditional drugs of abuse may serve as indicators of potential exposure to toxic adulterating substances during gestation. While there is cause for concern with respect to any unintentional drug exposure, illicit drug use during pregnancy, including uncontrolled dosing, poly-adulterant consumption, and the interactions of these drug mixtures, produces a significant public health threat to the neonate which warrants further study.
PubMed: 37025298
DOI: 10.3389/fped.2023.1127020 -
Molecules (Basel, Switzerland) Jan 2023Benzisothiazolinone (BIT), a biocide widely used as a preservative in household cleaning and personal care products, is cytotoxic to lung cells and a known skin allergen...
Benzisothiazolinone (BIT), a biocide widely used as a preservative in household cleaning and personal care products, is cytotoxic to lung cells and a known skin allergen in humans, which highlights the importance of assessing its toxicity and pharmacokinetics. In this study, a simple, sensitive, and accurate LC−MS/MS method for the quantification of BIT in rat plasma, urine, or tissue homogenates (50 μL) using phenacetin as an internal standard was developed and validated. Samples were extracted with ethyl acetate and separated using a Kinetex phenyl−hexyl column (100 × 2.1 mm, 2.6 μm) with isocratic 0.1% formic acid in methanol and distilled water over a run time of 6 min. Positive electrospray ionization with multiple reaction monitoring transitions of m/z 152.2 > 134.1 for BIT and 180.2 > 110.1 for phenacetin was used for quantification. This assay achieved good linearity in the calibration ranges of 2−2000 ng/mL (plasma and urine) and 10−1000 ng/mL (tissue homogenates), with r ≥ 0.9929. All validation parameters met the acceptance criteria. BIT pharmacokinetics was evaluated via an intravenous and dermal application. This is the first study that evaluated BIT pharmacokinetics in rats, providing insights into the relationship between BIT exposure and toxicity and a basis for future risk assessment studies in humans.
Topics: Humans; Rats; Animals; Chromatography, Liquid; Tandem Mass Spectrometry; Disinfectants; Phenacetin; Reproducibility of Results
PubMed: 36677902
DOI: 10.3390/molecules28020845 -
Pharmaceutics Dec 2022The solubility of active pharmaceutical ingredients is a mandatory physicochemical characteristic in pharmaceutical practice. However, the number of potential solvents...
Solubility Characteristics of Acetaminophen and Phenacetin in Binary Mixtures of Aqueous Organic Solvents: Experimental and Deep Machine Learning Screening of Green Dissolution Media.
The solubility of active pharmaceutical ingredients is a mandatory physicochemical characteristic in pharmaceutical practice. However, the number of potential solvents and their mixtures prevents direct measurements of all possible combinations for finding environmentally friendly, operational and cost-effective solubilizers. That is why support from theoretical screening seems to be valuable. Here, a collection of acetaminophen and phenacetin solubility data in neat and binary solvent mixtures was used for the development of a nonlinear deep machine learning model using new intuitive molecular descriptors derived from COSMO-RS computations. The literature dataset was augmented with results of new measurements in aqueous binary mixtures of 4-formylmorpholine, DMSO and DMF. The solubility values back-computed with the developed ensemble of neural networks are in perfect agreement with the experimental data, which enables the extensive screening of many combinations of solvents not studied experimentally within the applicability domain of the trained model. The final predictions were presented not only in the form of the set of optimal hyperparameters but also in a more intuitive way by the set of parameters of the Jouyban-Acree equation often used in the co-solvency domain. This new and effective approach is easily extendible to other systems, enabling the fast and reliable selection of candidates for new solvents and directing the experimental solubility screening of active pharmaceutical ingredients.
PubMed: 36559321
DOI: 10.3390/pharmaceutics14122828 -
Pharmaceutics Dec 2022Obtusifolin, a major anthraquinone component present in the seeds of Cassia tora, exhibits several biological activities, including the amelioration of memory...
Obtusifolin, a major anthraquinone component present in the seeds of Cassia tora, exhibits several biological activities, including the amelioration of memory impairment, prevention of breast cancer metastasis, and reduction of cartilage damage in osteoarthritis. We aimed to evaluate the inhibitory effects of obtusifolin and its analogs on CYP1A enzymes, which are responsible for activating procarcinogens, and investigate its inhibitory mechanism and chemopreventive effects. P450-selective substrates were incubated with human liver microsomes (HLMs) or recombinant CYP1A1 and CYP1A2 in the presence of obtusifolin and its four analogs. After incubation, the samples were analyzed using liquid chromatography-tandem mass spectrometry. Molecular docking simulations were performed using the crystal structure of CYP1A2 to identify the critical interactions between anthraquinones and human CYP1A2. Obtusifolin potently and selectively inhibited CYP1A2-mediated phenacetin O-deethylation (POD) with a Ki value of 0.031 µM in a competitive inhibitory manner in HLMs, whereas it exhibited negligible inhibitory effect against other P450s (IC50 > 28.6 µM). Obtusifolin also inhibited CYP1A1- and CYP1A2-mediated POD and ethoxyresorufin O-deethylation with IC50 values of <0.57 µM when using recombinant enzymes. Our molecular docking models suggested that the high CYP1A2 inhibitory activity of obtusifolin may be attributed to the combination of hydrophobic interactions and hydrogen bonding. This is the first report of selective and potent inhibitory effects of obtusifolin against CYP1A, indicating their potential chemopreventive effects.
PubMed: 36559174
DOI: 10.3390/pharmaceutics14122683 -
Frontiers in Microbiology 2022Mycotoxins, fungal secondary metabolites, are ubiquitously present in food commodities. Acute exposure to high levels or chronic exposure to low levels has an impact on...
Unravelling the pharmacokinetics of aflatoxin B1: determination of Michaelis-Menten constants, intrinsic clearance and the metabolic contribution of CYP1A2 and CYP3A4 in pooled human liver microsomes.
Mycotoxins, fungal secondary metabolites, are ubiquitously present in food commodities. Acute exposure to high levels or chronic exposure to low levels has an impact on the human body. The phase I metabolism in the human liver, performed by cytochrome P450 (CYP450) enzymes, is accountable for more than 80% of the overall metabolism of exogenous and endogenous compounds. Mycotoxins are (partially) metabolized by CYP450 enzymes. In this study, research was performed on CYP450 probes and aflatoxin B1 (AFB1), a carcinogenic mycotoxin, to obtain pharmacokinetic data on AFB1, required for further experimental work. The CYP450 probes of choice were a CYP3A4 substrate, midazolam (MDZ) and a CYP1A2 substrate, phenacetin (PH) since these are the main metabolizing phase I enzymes of AFB1. Linearity experiments were performed on the three substrates indicating that linear conditions were achieved at a microsomal protein concentration and incubation time of 0.25 mg/ml and 5 min, 0.50 mg/ml and 20 min and 0.25 mg/ml and 5 min for MDZ, PH and AFB1, respectively. The was determined in human liver microsomes and was estimated at 2.15 μM for MDZ, 40.0 μM for PH and 40.9 μM for AFB1. The associated values were 956 pmol/(mg.min) (MDZ), 856 pmol/(mg.min) (PH) and 11,536 pmol/(mg.min) (AFB1). Recombinant CYP systems were used to determine CYP450-specific Michaelis-Menten values for AFB1, leading to a CYP3A4 of 49.6 μM and an intersystem extrapolation factor (ISEF) corrected of 43.6 pmol/min/pmol P450 and a CYP1A2 of 58.2 μM and an ISEF corrected of 283 pmol/min/pmol P450. An activity adjustment factor (AAF) was calculated to account for differences between microsome batches and was used as a correction factor in the determination of the human hepatic clearance for MDZ, PH and AFB1. The hepatic blood clearance corrected for the AAF CL, CL CL and CL were determined in HLM at 44.1 L/h, 21.7 L/h, 40.0 L/h and 38.5 L/h. Finally, inhibition assays in HLM showed that 45% of the AFB1 metabolism was performed by CYP3A4/3A5 enzymes and 49% by CYP1A2 enzymes.
PubMed: 36110298
DOI: 10.3389/fmicb.2022.988083 -
MethodsX 2022The data presented in this article are related to the research article entitled "Cytochrome P450 inhibition activities of non-standardized botanical products" [1], in...
The data presented in this article are related to the research article entitled "Cytochrome P450 inhibition activities of non-standardized botanical products" [1], in which the possible CYP inhibitory properties of botanical products were investigated. This article describes the optimization and bioanalytical method validation of the CYP (Cytochrome P450 inhibition assay) inhibition assays, namely, phenacetin O-deethylase assay, testosterone 6β-hydroxylase assay, felodipine dehydrogenase assay and midazolam 1'-hydroxylase assay using LC-MS/MS.
PubMed: 36081487
DOI: 10.1016/j.mex.2022.101827 -
Sheng Wu Yi Xue Gong Cheng Xue Za Zhi =... Aug 2022Primary human hepatocytes (PHH) are the gold standard of human liver model for drug screening. However, a problem of culturing PHH is the rapid decline of cytochrome...
Primary human hepatocytes (PHH) are the gold standard of human liver model for drug screening. However, a problem of culturing PHH is the rapid decline of cytochrome P450 (CYP450) activity, which plays an important role in drug metabolism. In this study, thermo-responsive culture dishes were used to explore the conditions for murine embryonic 3T3-J2 fibroblasts to form cell sheet. Based on the cell sheet engineering technology, a three-dimensional (3D) "sandwich" co-culture system of 3T3-J2 cell sheet/PHH/collagen gel was constructed. The tissue structure and protein expression of the model section were observed by hematoxylin eosin staining and immunofluorescence staining respectively. Phenacetin and bupropion were used as substrates to determine the activity of CYP450. The contents of albumin and urea in the system were determined by enzyme linked immunosorbent assay (ELISA). The results showed that the complete 3T3-J2 cell sheet could be obtained when the cell seeding density was 1.5×10 /dish (35 mm dish) and the incubation time at low temperature was 60 min. Through cell sheet stacking, a 3D liver model was developed. Compared with the two-dimensional (2D) model, in the 3D model, the cell-cell and cell-matrix connections were tighter, the activities of cytochrome P450 CYP1A2 and cytochrome P450 CYP2B6 were significantly increased, and the secretion levels of albumin and urea were increased. These indexes could be maintained stably for 21 d. Therefore, cell sheet stacking is helpful to improve the level of liver function of 3D liver model. This model is expected to be used to predict the metabolism of low-clearance drugs in preclinical, which is of great significance for drug evaluation and other studies.
Topics: Albumins; Animals; Cytochrome P-450 Enzyme System; Hepatocytes; Humans; Liver; Mice; Urea
PubMed: 36008342
DOI: 10.7507/1001-5515.202108056 -
Annals of Translational Medicine May 2022Tea, the world's second most popular drink, is an essential part of some people's lives. Thus, this study aimed to explore potential tea-drug interactions with a view to...
BACKGROUND
Tea, the world's second most popular drink, is an essential part of some people's lives. Thus, this study aimed to explore potential tea-drug interactions with a view to promoting the rational administration of drugs.
METHODS
A specific and sensitive approach involving high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) and a probe cocktail was established and validated to evaluate the inhibitory effects of four teas on five cytochrome P450 (CYP450) enzymes in rats. Metoprolol tartrate (MT), omeprazole (OMP), phenacetin (PNT), tolbutamide (TOL), and testosterone (T) were selected as the probe drugs for CYP2D6, CYP2C19, CYP1A2, CYP2C6, and CYP3A1/2, respectively, and were simultaneously quantified in the multiple reaction monitoring (MRM) mode with positive electrospray ionization (ESI+) in a single 12-min run.
RESULTS
The extraction recoveries, matrix effect values, as well as intra/interday accuracy and precision met the determination standards. CYP1A2 and CYP2C6 were strongly inhibited by green tea, and CYP2C6 was also strongly inhibited by Pu'er tea. Ti Kuan Yin tea had a weak inhibitory effect, and black tea had only a slight inhibitory effect, on CYP1A2. Furthermore, the four types of tea did not have significantly altered the activity of CYP2D6, CYP2C19, and CYP3A1/2 .
CONCLUSIONS
The method used in the present study was successfully applied to assess the inhibitory effects of aqueous extracts of four types of tea on CYP450 isoforms . The results suggest that different types of tea have different effects on drug metabolism.
PubMed: 35928758
DOI: 10.21037/atm-21-5490