Combinatorial Chemistry & High Throughput Screening - Volume 23, Issue 10, 2020

Aim: A new, accurate and sensitive reversed-phase high-performance liquid chromatography (RP-HPLC) as an analytical method for the quantitative determination of eight antifungal drugs in spiked human plasma has been described optimized and validated. Materials and Methods: The analyzed compounds were voriconazole (VOR), luliconazole (LUL), clotrimazole (CLO), tioconazole (TIO), posaconazole (POS), ketoconazole (KET), sertaconazole (SER) and terconazole (TER). Results: The separation of the analyzed compounds was conducted using a novel pentabromobenzyl column known as COSMOSIL PBB-R (150 mm × 4.6 mm I.D., particle size 5 μm). The analysis of the studied drugs was determined within 14 min using a diode array detector and the mobile phase consisted of: 10 mM potassium dihydrogen phosphate buffer (pH 2.1): Methanol (2: 98 v/v). A linear response was observed for all compounds in the range of concentration studied. Sample preparation was done through liquid-liquid extraction using diethyl ether. Conclusion: This proposed method was validated in terms of linearity, limit of quantification, limit of detection, accuracy, precision and selectivity. The method was successfully applied for the determination of these drugs in their pharmaceutical formulations and in human plasma samples.

Aims and Objective: The development of easy, accurate, reliable technique which is characterized by low cost, minimal sample pre-treatment, and short analysis time to monitor RFX residues in milk samples before distribution to consumers. Background: Literature survey reveals several analytical methods, including high-performance liquid chromatography (HPLC), ultra-performance liquid chromatography (UPLC) and thin-layer chromatography (TLC)-densitometry. These methods are time consuming, require additional steps like preconcentration or multisolvent extraction, trained technicians, and expensive instruments. Materials and Methods: The electrochemical analysis of RFX was effectively established by the adsorptive stripping method on GCE due to the effective interfacial accumulation of RFX on the electrode surface. The RFX adsorptive accumulation is followed by electrochemical measurement of the accumulated analyte. Results: The electrochemical oxidation of RFX was studied at glassy carbon electrodes (GCE) in Britton-Robinson buffer (BR) solutions over the pH range from 2.0-12.0 using cyclic and differential pulse voltammetry (DPV). The oxidation of the drug was accomplished in a single irreversible, adsorption-controlled step within the pH range 4.0-9.0. Therefore, the application of GCE for a sensitive and selective quantification of RFX by adsorptive stripping voltammetry was reported. This format was satisfactorily applied for the determination of RFX in bovine milk. Limit of detection (LOD) of 1.25 μg kg-1 of milk and mean recoveries of 97.8 to 107.5% were achieved. Conclusion: The proposed method might be competitive with the HPLC techniques. The detection limit found for RFX on GCE for milk samples, after medium exchange, was well below the MRLs, the maximum concentration of a veterinary drug residue legally permissible in food, are proposed by the European Medicines Agency.

Aim and Scope: Due to the serious toxicological risks and their widespread use, quantitative determination of phthalates in cosmetic products have importance for public health. The aim of this study was to develop a validated simple, rapid and reliable high-performance liquid chromatography (HPLC) method for the determination of phthalates which are; dimethyl phthalate (DMP), diethyl phthalate (DEP), benzyl butyl phthalate (BBP), di-n-butyl phthalate (DBP), di(2- ethylhexyl) phthalate (DEHP), in cosmetic products and to investigate these phthalate (PHT) levels in 48 cosmetic products marketing in Sivas, Turkey. Materials and Methods: Separation was achieved by a reverse-phase ACE-5 C18 column (4.6 x 250 mm, 5.0 μm). As the mobile phase, 5 mM KH2PO4 and acetonitrile were used gradiently at 1.5 ml min-1. All PHT esters were detected at 230 nm and the run time was taking 21 minutes. Results: This method showed the high sensitivity value the limit of quantification (LOQ) values for which are below 0.64 μg mL^-1 of all phthalates. Method linearity was 137;¥0.999 (r²). Accuracy and precision values of all phthalates were calculated between (-6.5) and 6.6 (RE%) and 137;¤6.2 (RSD%), respectively. Average recovery was between 94.8% and 99.6%. Forty-eight samples used for both babies and adults were successfully analyzed by the developed method. Results have shown that, DMP (340.7 μg mL^-1 ±323.7), DEP (1852.1 μg mL^-1 ± 2192.0), and DBP (691.3 μg mL^-1 ± 1378.5) were used highly in nail polish, fragrance and cream products, respectively. Conclusion: Phthalate esters, which are mostly detected in the content of fragrance, cream and nail polish products and our research in general, are DEP (1852.1 μg mL^-1 ± 2192.0), DBP (691.3 μg mL^-1 ± 1378.5) and DMP (340.7 μg mL^-1 ±323.7), respectively. Phthalates were found in the content of all 48 cosmetic products examined, and the most detected phthalates in general average were DEP (581.7 μg mL^-1 + 1405.2) with a rate of 79.2%. The unexpectedly high phthalate content in the examined cosmetic products revealed a great risk of these products on human health. The developed method is a simple, sensitive, reliable and economical alternative for the determination of phthalates in the content of cosmetic products, it can be used to identify phthalate esters in different products after some modifications.

Aim and Objective: Methyldopa is one of the medications that is used for the treatment of hypertension. Therefore, the determination of methyldopa in the presence of other biological components is essential. In this work, a promising electrochemical sensor based on CoFe2O4 magnetic nanoparticles modified glassy carbon electrode (CoFe2O4/GCE) was developed for electrochemical determination of methyldopa in the presence of uric acid. Cobalt ferrite nanoparticles were synthesized via chemical method. Materials and Methods: Characterizing the CoFe2O4 was investigated by field emission scanning electron microscopy (FESEM), X-ray diffraction (XRD), energy dispersive X-ray spectroscopy (EDX), transmission electron microscope (TEM), and cyclic voltammetry techniques. Results: Under the optimal experimental conditions, the current response of the electrochemical sensor obtained with differential pulse voltammetry was increased linearly in the concentration range from 1.45 to 15.1 μmol L^-1 with the detection limit of 1.07 μmol L^-1 for methyldopa. Also, by using the proposed method, methyldopa and uric acid could be analyzed in a mixture independently. The difference in peak potential for analytes is about 150 mV. Conclusion: The present sensor was successfully applied for the determination of methyldopa in the presence of uric acid in biological samples and the pharmaceutical samples with satisfactory results.

Aims and Objective: The plasma level of mirtazapine (MIR) varies between individuals primarily depending on the differences in metabolism during pharmacotherapy. CYP2D6 takes the role as a major enzyme in MIR metabolism and POR enzyme donates an electron to CYP2D6 for its activity. Single nucleotide polymorphisms in the genes encoding pharmacokinetic enzymes may cause changes in enzyme activity, leading to differences in metabolism of the drug. Our aim was to assess the influence of CYP2D6*4 and POR*28 polymorphisms on MIR plasma levels in Turkish psychiatric patients. Materials and Methods: The association between genetic variations and plasma level of MIR was investigated on 54 patients. CYP2D6*4 and POR*28 polymorphisms were analysed using Polymerase Chain Reaction- Restriction Fragment Length Polymorphism (PCR-RFLP) and plasma MIR levels were measured using HPLC. Results: Allele frequencies of CYP2D6*4 and POR*28 were 0.11 and 0.39, respectively in the study population. The results showed that CYP2D6*4 allele carriers have higher C/D MIR levels while POR*28 allele carriers have lower C/D MIR levels. Combined genotype analyses also revealed that individuals with CYP2D6*1/*1 - POR*28/*28 genotype have a statistically lower C/D MIR level (0.95 ng/ml/dose) when compared with individuals with CYP2D6*1/*1 - POR*1/*1 genotype (1.52 ng/ml/dose). Conclusion: Our results indicate that CYP2D6*4 and POR*28 polymorphisms may have a potential in the explanation of differences in plasma levels in MIR treated psychiatric patients. A combination of these variations may be beneficial in increasing drug response and decreasing adverse drug reactions in MIR psychopharmacotherapy.

Aim and Objective: Parkinson’s disease (PD) is the second most common neurodegenerative disease. It is a multifactorial disorder (caused by aging, environmental, and genetic factors). Metabolomics can help explore the biomarker profiles for aging. Recent studies showed an association between the glutathione S-transferases (GSTs) polymorphisms and PD risk. The purpose of this study was to evaluate the association of this genetic polymorphism and the metabolomic profile in PD Tunisian patients, in order to identify effective biomarkers in the genetic differentiation. Materials and Methods: In this study, the metabolomic profile changes related to GSTs polymorphism were searched in 54 Tunisian PD patients treated with L-dopa, using a gas chromatography-mass spectrometry (GC-MS) technique. Results: The study results showed that mannose, methyl stearate, and three other unknown metabolites, increased in patients with GSTM1 positive genotype, while glycolic acid, porphine, monomethyl phosphate, fumaric acid, and three other unknown metabolites decreased in patients with GSTM1 positive genotype. Subsequently, the levels of glycolic acid, erythronic acid, lactic acid, citric acid, fructose, stearic acid, 2-amino-2-methyl-1,3-propanediol and three other unknown metabolites increased in patients with GSTM1 positive genotype, while the levels of proline, valine and two unknown metabolites decreased with GSTT1 positive genotype. Conclusion: All these altered metabolites are related to energy metabolism and it can be concluded that GSTs polymorphism based the shifting in energy metabolism and led to oxidative stress.

Objective: Literature study revealed the poor mechanical strength of chitosan-based microparticles. Our research aimed at developing sufficient strength of microparticle with a suitable concentration of chitosan and non-ionic surfactants such as poloxamer-188 (pluronic). It also aimed to develop and study the effect of variables for prepared microparticles utilizing insilico screening methodology, such as reduced factorial design, followed by optimization. Methods: Preliminary trial batches were prepared with variable concentration of chitosan and poloxamer-188 utilizing cross-linked ion-gelation technique. A 20% w/v sodium citrate solution was used as a cross-linking solution. The resolution-IV of 24-1 reduced factorial design was selected to screen the possible and significant independent variables or factors in the dosage form design. A total number of eight runs were suggested by statistical software and responses were recorded. The responses such as spreadability, pH, viscosity and percentage of drug released at 12 h were considered in the screening study. Based on the result, selected factors were included in the optimization technique, including graphical and numerical methods. Results: The signified factors based on reduced two-level factorial screening design with randomized subtype, were identified by Half-normal and Pareto chart. Mathematical fitting and analysis were performed by the factorial equation during the optimization process. The validation and fitting of models were suggested and evaluated by p-value, adjusted R2, and predicted R2 values. The significant and non-significant terms were evaluated, followed by finding the optimal concentration and region with yellow color highlighted in an overlay plot. Based on the data obtained by the overlay study, the final formulation batch was prepared and the observed value was found to be pretty much nearer as compared to predicted values. Drug-polymer interaction study included attenuated total reflectance, differential scanning calorimetry, and X-Ray diffraction study. Conclusion: The principal of the study design was based on finding the prefixed set parameter values utilizing the concept of in-silico screening technique and optimization with a minimal number of trials and study expenses. It concluded that Poloxamer-188 (0.94%), chitosan (2.38%), swelling time (1.81 h), and parts of chitosan (78.51%) in a formulation batch would fulfill the predetermined parameter with specific values.

Aims: To prepare lamivudine (LAM)-loaded-nanoparticles (NPs) that can be used in lung cancer treatment. To change the antiviral indication of LAM to anticancer. Background: The development of anticancer drugs is a difficult process. One approach to accelerate the availability of drugs is to reclassify drugs approved for other conditions as anticancer. The most common route of administration of anticancer drugs is intravenous injection. Oral administration of anticancer drugs may considerably change current treatment modalities of chemotherapy and improve the life quality of cancer patients. There is also a potentially significant economic advantage. Objective: To characterize the LAM-loaded-NPs and examine the anticancer activity. Methods: LAM-loaded-NPs were prepared using Nano Spray-Dryer. Properties of NPs were elucidated by particle size (PS), polydispersity index (PDI), zeta potential (ZP), SEM, encapsulation efficiency (EE%), dissolution, release kinetics, DSC and FT-IR. Then, the anticancer activity of all NPs was examined. Results: The PS values of the LAM-loaded-NPs were between 373 and 486 nm. All NPs prepared have spherical structure and positive ZP. EE% was in a range of 61-79%. NPs showed prolonged release and the release kinetics fitted to the Weibull model. NPs structures were clarified by DSC and FT-IR analysis. The results showed that the properties of NPs were directly related to the drug:polymer ratio of feed solution. NPs have potential anticancer properties against A549 cell line at low concentrations and non-toxic to CCD 19-Lu cell line. Conclusion: NPs have potential anticancer properties against human lung adenocarcinoma cells and may induce cell death effectively and be a potent modality to treat this type of cancer. These experiments also indicate that our formulations are non-toxic to normal cells. It is clear that this study would bring a new perspective to cancer therapy.

Aim and Objective: Melatonin is an essential biomarker for sleep-related disorders. Reliable methods of analysis are needed for melatonin. Therefore, a new chromophore (Rhodamine B) was proposed for the assay of melatonin; this method succeeded to enlarge the working concentration range and to decrease the limit of determination comparing with the method that just used the native fluorescence of melatonin. Materials and Methods: Rhodamine B was proposed as a new chromophore for the assay of melatonin in biological, food, and pharmaceutical samples. Fluorescence was used for the determination of melatonin. Results: The results obtained using Rhodamine B were compared with those obtained by the native fluorescence of melatonin. Using the new chromophore, melatonin was determined in the concentration range between 0.01 and 50 pmol L-1, with the detection limit of 2.4 fmol L-1. The recovery of melatonin was higher than 98.00% with a relative standard deviation of less than 0.10%, when the method was applied for the assay of melatonin in samples such as breast milk, whole blood, milk powder, and pharmaceutical formulations. Conclusion: Utilization of Rhodamine B enlarged the linear concentration range for the assay of melatonin and decreased the detection limit, making possible the assay of melatonin in a variety of samples such as pharmaceuticals, food, and biological samples.

Background: Allergic rhinitis, acute nasal congestion and sinusitis are one of the most common health problems and have a major effect on the quality of life. Several medications are used to improve the symptoms of such diseases in humans. Pharmaceutical pomade form containing Ephedrine (EPD) HCl, Naphazoline (NPZ) HCl, Antazoline (ANT) HCl, and Chlorobutanol (CLO) is one of them. Objective: For these reasons, this study includes the development of spectrophotometric and chromatographic methods for the determination of EPD HCl, NPZ HCl, ANT HCl, and CLO active agents in the pharmaceutical pomade. Method: In the spectrophotometric method, third-order derivative of the amplitudes at 218 nm n=5 and the first-order derivative of the amplitudes 254 nm n=13 was selected for the determination of EPD, ANT, respectively while NPZ was determined by the second derivative at 234 nm and n=21. Colorimetric detection was applied for assay analysis of CLO at 540 nm. Furthermore, a reverse phase high performance liquid chromatographic (RP- HPLC) method has been developed and optimized by using Agilent Zorbax Eclipse XDB C18 (75 mm x 3.0 mm, 3.5μm) column. The column temperature was 40°C, binary gradient elution was used and the mobile phase consisted of 15 mM phosphate buffer in distilled water (pH 3.0) and methanol, and the flow rate was 0.6 mL min^-1and the UV detector was detected at 210 nm. The linear operating range was obtained as 11.97-70, 0.59-3, 2.79-30, and 2.92-200 μg mL^-1 for EPD HCl, NPZ HCl, ANT HCl, and CLO respectively. Results: The LOD values were found to be 3.95, 0.19, 0.92 and 0.96 μg mL^-1 for EPD HCl, NPZ HCl, ANT HCl, and CLO in the spectrophotometric method, respectively. The linear ranges in the RP-HPLC method were 8.2-24.36 μg mL^-1, 0.083 - 0.75 μg/mL, 2.01-7.5 μg mL^-1 and 2.89-24.4 μg mL^-1 for EPD HCl, NPZ HCl, ANT HCl, and CLO, respectively. The LOD values in the validation studies were 2.7, 0.025, 0.66 and 0.86 μg mL^-1 for EPD HCl, NPZ HCl, ANT HCl, and CLO in RP-HPLC method respectively. Conclusion: The results of the spectrophotometric and chromatographic methods were compared and no differences were found between the two methods.

Aim and Objective: Myocardial infarction, cerebral infarction, and other diseases caused by vascular obstruction have always jeopardized human life and health. Several reports indicate that Rhei Radix et Rhizoma has a good clinical effect in the prevention and treatment of cardiovascular diseases. Owing to the complexity of herbal medicine, the pharmacodynamic mechanism of Rhei Radix et Rhizoma is still unclear. The objectives of this study were to explore the two-way adjustment mechanism of Rhei Radix et Rhizoma and provide a new solution for the prevention and treatment of cardiovascular disease. Materials and Methods: This study used data mining, reverse pharmacophore matching, network construction, GO and KEGG Analysis, and molecular docking to investigate the two-way adjustment mechanism of Rhei Radix et Rhizoma. The methods used were based on systems pharmacology and big data analysis technology. Results: The results suggest that Rhei Radix et Rhizoma uses a two-way adjustment of activating blood circulation, as well as blood coagulation in the prevention and treatment of cardiovascular diseases. The components involved in activating blood circulation are mainly anthraquinone components. The corresponding targets are NOS2, NOS3, CALM1, and the corresponding pathways are calcium signaling pathway, VEGF signaling pathway, platelet activation, and the PI3K-Akt signaling pathway. For blood coagulation, the components are mainly tannin components; the corresponding targets are F2, F10, ELANE, and the corresponding pathways are the neuroactive ligand-receptor interaction, complement and coagulation cascades. Conclusion: This study indicated that Rhei Radix et Rhizoma exerts the two-way adjustment of activating blood circulation and blood coagulation in the prevention and treatment of cardiovascular diseases. It can make up for the side effects of the existing blood circulation drugs for cardiovascular disease, only activating blood circulation, and the uncontrollable large-area bleeding due to the long-term use of the drugs. This study provides a material basis for the development of new blood-activating drugs based on natural medicine.

Background: Several medicinal plants are being used in Indian medicine systems from ancient times. However, in most cases, the specific molecules or the active ingredients responsible for the medicinal or therapeutic properties are not yet known. Objective: This study aimed to report a computational protocol as well as a tool for generating novel potential drug candidates from the bioactive molecules of Indian medicinal and aromatic plants through the chemoinformatics approach. Methods: We built a database of the Indian medicinal and aromatic plants coupled with associated information (plant families, plant parts used for the medicinal purpose, structural information, therapeutic properties, etc.) We also developed a Java-based chemoinformatics open-source tool called DoMINE (Database of Medicinally Important Natural products from plantaE) for the generation of virtual library and screening of novel molecules from known medicinal plant molecules. We employed chemoinformatics approaches to in-silico screened metabolites from 104 Indian medicinal and aromatic plants and designed novel drug-like bioactive molecules. For this purpose, 1665 ring containing molecules were identified by text mining of literature related to the medicinal plant species, which were later used to extract 209 molecular scaffolds. Different scaffolds were further used to build a focused virtual library. Virtual screening was performed with cluster analysis to predict drug-like and lead-like molecules from these plant molecules in the context of drug discovery. The predicted drug-like and lead-like molecules were evaluated using chemoinformatics approaches and statistical parameters, and only the most significant molecules were proposed as the candidate molecules to develop new drugs. Results and Conclusion: The supra network of molecules and scaffolds identifies the relationship between the plant molecules and drugs. Cluster analysis of virtual library molecules showed that novel molecules had more pharmacophoric properties than toxicophoric and chemophoric properties. We also developed the DoMINE toolkit for the advancement of natural product-based drug discovery through chemoinformatics approaches. This study will be useful in developing new drug molecules from the known medicinal plant molecules. Hence, this work will encourage experimental organic chemists to synthesize these molecules based on the predicted values. These synthesized molecules need to be subjected to biological screening to identify potential molecules for drug discovery research.